T cell epitopes of the major allergens from dermatophagoides (house dust mite)

ABSTRACT

The present invention provides isolated peptides of the major protein allergens of the genus Dermatophagoides. Peptides within the scope of the invention comprises at least one T cell epitope, or preferably at least two T cell epitopes of a protein allergen selected from the allergens Der p I, Der p II, Der f I, or Der f II. The invention also pertains to modified peptides having similar or enhanced therapeutic properties as the corresponding, naturally-occurring allergen or portion thereof, but having reduced side effects. The invention further provides nucleic acid sequences coding for peptides of the invention. Methods of treatment or of diagnosis of sensitivity to house dust mites in an individual and therapeutic compositions comprising one or more peptides of the invention are also provided.

RELATED APPLICATIONS

This application is a divisional of U.S. Ser. No. 08/445,307, filed May19, 1995 which is a continuation-in-part of U.S. Ser. No. 08/227,772filed Apr. 14, 1994, now abandoned, which is a continuation-in-part ofPCT/US93/03471 filed Apr. 14, 1993.

BACKGROUND OF THE INVENTION

Recent reports have documented the importance of responses to the GroupI (e.g., Der p I and Der f I) and Group II (e.g., Der p II and Der f II)protein allergens in house dust mite allergy. For example, it has beendocumented that over 60% of patients have at least 50% of theiranti-mite antibodies directed towards these proteins (e.g., Lind, P. etal., Allergy, 39:259-274 (1984); van der Zee, J. S. et al., JournalAllergy and Clinical Immunology, 81:884-896 (1988)). It is possible thatchildren show a greater degree of reactivity to the Group I and Group IIallergens (Thompson, P. J., et al., Immunology, 64:301-314 (1988)).Allergy to mites of the genus Dermatophagoides (D.) is associated withconditions such as asthma, rhinitis and ectopic dermatitis. Two species,D. pteronyssinus and D. farinae, predominate and, as a result,considerable effort has been expended in trying to identify theallergens produced by these two species.

A concerted effort has been made to characterize by gene cloning themajor allergens from both D. pteronyssinus and D. farinae. Consequently,several publications have reported the complete nucleotide sequences ofseveral allergens including Der p I (Thomas, W. R., et al.,International Archives of Allergy and Applied Immunology. 85:127-129(1988); and Chua, K. Y., et al., Journal of Experimental Medicine,167:175-182 (1988)), Der p II (Chua, K. Y., et al., InternationalArchives of Allergy and Applied Immunology, 91:118-123 (1990)), Der f I(Dilworth, R. J., et al., Clinical and Experimental Allergy, 21:25-32(1891)), Der f II (Yuuki, T., et al., Japan Journal Allergol.,39:557-461 (1990); and Trudinger, M., et al., Clinical and ExperimentalAllergy, 21:33-37 (1991)) and a low molecular weight allergen (Ovey, E.R., et al., Journal of Experimental Medicine, 170:1457-1462 (1989)).

The published nucleotide sequences of cDNAs encoding Der p I and Der f Idemonstrate that these two proteins are highly homologous at the aminoacid level (81% identity) and that the mature protein products arecomprised of 222 and 223 residues, respectively (Chua, K. Y., et al.,Journal of Experimental Medicine, 167:175-182 (1988); and Dilworth, R.J., et al., supra)). The protein allergens Der p II and Der f II areboth comprised of 129 residues, and are also highly homologous (88%identity) in amino acid sequence (Trudinger, M., et al. supra; Yuuki,T., et al. supra); Chua, K. Y., et al, International Archives of Allergyand Applied Immunology, 91:118-123 (1990)).

The isolation of cDNAs clones encoding Der p I and Der p II haspermitted antibody binding studies on the recombinant antigens (Green,W. K., et al., International Archives of Allergy and Applied Immunology,92:30-38 (1990); Chua, K. Y., et al., International Archives of Allergyand Applied Immunology, 91:124-129 (1990)). Complementary DNA fragmentsof Der p I have been expressed in E. coli and IgE binding studies withpooled human mite allergic IgE sera have demonstrated binding andnon-binding regions throughout the molecule (Thomas, W. R., et al., In:Epitopes of Atopic Allergens, Proceedings of Workshop from XIV Congressof the European Academy of Allergy and Clinical Immunology, Berlin,September 1989. pp 77-82). T cell epitopes of Der p I have been reported(O'Hehir, R. E., et al., Annual Review Immunology, 9:67-95 (1991);Stewart, G. A., et al., In: Epitopes of Atopic Allergens, Proceedings ofWorkshop from XIV Congress of the European Academy of Allergy andClinical Immunology, Berlin, September 1989. pp 41-47; Yessel, H., etal., In: T cell Activation in Health and Disease: Discrimination BetweenImmunity and Tolerance, Conference 22-26 Sep. 1990, Trinity College,Oxford, U.K. and Hessel, H., et al., Journal of Immunology, 148(3):738-745 (Feb. 1, 1992).

SUMMARY OF THE INVENTION

The present invention provides isolated peptides of the major proteinallergens of the genus Dermatophagoides. Preferred peptides within thescope of the invention comprise at least one T cell epitope, and maycomprise at least two T cell epitopes of a protein allergen selectedfrom the allergens Der p I, Der p II, Der f I, or Der f II. Theinvention further provides peptides comprising at least two regions,each region comprising at least one T cell epitope of a mite proteinallergen. The regions are derived from the same or from differentprotein allergens of the genus Dermatophagoides.

The invention also provides modified peptides having similar or enhancedtherapeutic properties as the corresponding, naturally-occurringallergen or portion thereof, but having reduced side effects, as well asmodified peptides having improved properties such as increasedsolubility and stability. Peptides of the invention are capable ofmodifying, in a house dust mite-sensitive individual to whom they areadministered, the allergic response of the individual to a house dustmite allergen or an allergen immunologically cross-reactive with housedust mite allergen. Methods of treatment or of diagnosis of sensitivityto house dust mite in an individual and therapeutic compositionscomprising one or more peptides of the invention are also provided.

The present invention further provides optimized therapeuticcompositions and multipeptide formulations comprising "unique" peptidesof the invention. Such therapeutic compositions have been optimized toaccomodate and maintain the unique characteristics of the "unique"peptides of the invention, and at the same time provide maximumtherapeutic effect when used in therapeutic regimens for the treatmentof house dust mite allergy in humans.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the subcloning and expression of Group I and Group IIallergens from D. pteronyssinus and D. farinae.

FIG. 2a shows adaptors used in the expression of Der p I and Der f I andFIG. 2b shows primers for amplification of Der f II and Der p II and aDer f II mutagenesis primer.

FIG. 3 shows various peptides of desired lengths derived from the Der pI and Der p II protein allergens.

FIG. 4 shows various peptides of desired lengths derived from the Der fI and Der f II protein allergens.

FIG. 5 is a graphic representation depicting the responses of T celllines from 33 patients primed in vitro to either purified native (N) orrecombinant (R) Der p I protein and analyzed for response to variousoverlapping Der p I peptides by percent of responses with a T cellStimulation Index (S.I.) of at least 2 within the individuals tested,the mean T cell stimulation index of positive responses for the peptideand the ranked sum of peptide responses.

FIG. 6 is a graphic representation depicting the responses of T celllines from 16 patients primed in vitro to the Der f I protein andanalyzed for response to various overlapping Der f I peptides by percentof responses with an (S.I.) of at least 2 within the individuals tested,the mean T cell stimulation index of positive responses for the peptideand the ranked sum of peptide responses.

FIG. 7 is a graphic representation depicting the responses of T celllines from 14 patients primed in vitro to the Der p I protein andanalyzed for response to various overlapping Der p I peptides andsubstantially matching Der f I peptides by percent of responses with anS.I. of at least 2 within the individuals tested, the mean T cellstimulation index of positive responses for the peptide and the rankedsum of peptide responses.

FIG. 8 is a graphic representation depicting the responses of T celllines from 8 patients primed in vitro to the Der f I protein andanalyzed for response to various overlapping Der f I peptides andsubstantially matching Der p I peptides by percent of responses with anS.I. of at least 2 within the individuals tested, the mean T cellstimulation index of positive responses for the peptide and the rankedsum of peptide responses.

FIG. 9 is a graphic representation depicting the responses of T celllines from 29 patients primed in vitro to the Der p 11 protein andanalyzed for response to various overlapping Der p II peptides bypercent of responses with an S.I. of at least 2 within the individualstested, the mean T cell stimulation index of positive responses for thepeptide and the ranked sum of peptide responses.

FIG. 10 is a graphic representation depicting the responses of T celllines from 10 patients primed in vitro to the Der f II protein andanalyzed for response to various overlapping Der f II peptides bypercent of responses with an S.I. of at least 2 within the individualstested, the mean T cell stimulation index of positive responses for thepeptide and the ranked sum of peptide responses.

FIG. 11 is a graphic representation depicting the responses of T celllines from 10 patients primed in vitro to the Der f II protein andanalyzed for response to various overlapping Der f II peptides andsubstantially matching Der p II peptides by percent of responses with anS.I. of at least 2 within the individuals tested, the mean T cellstimulation index of positive responses for the peptide and the rankedsum of peptide responses.

FIG. 12 is a graphic representation depicting the responses of T celllines from 26 patients primed in vitro to the Der p II protein andanalyzed for response to various overlapping Der f II peptides bypercent of responses with an S.I. of at least 2 within the individualstested, the mean T cell stimulation index of positive responses for thepeptide and the ranked sum of peptide responses.

FIG. 13 is a graphic representation depicting the responses of T celllines from 33 patients primed in vitro to the Der p I protein andanalyzed for response to selected peptides of desired lengths derivedfrom the Der p I protein allergen by percent of responses with an S.I.of at least 2 within the individuals tested, the mean T cell stimulationindex of positive responses for the peptide and the ranked sum ofpeptide responses.

FIG. 14 is a graphic representation depicting the responses of T celllines from 9 patients primed in vitro to the Der f I protein andanalyzed for response to selected peptides of desired lengths derivedfrom the Der f I protein allergen, by percent of responses with an S.I.of at least 2 within the individuals tested and the mean T cellstimulation index of positive responses for the peptide and the rankedsum of peptide responses.

FIG. 15a is a graphic representation depicting the responses of T celllines from 30 matched patients primed in vitro to the Der p I proteinand analyzed for response to selected peptides of desired lengthsderived from the Der p I and the Der f I protein allergens, by percentof responses with an S.I. of at least 2 within the individuals testedand the mean T cell stimulation index of positive responses for thepeptide.

FIG. 15b is a graphic representation derived from the same data shown inFIG. 15a showing the response of Der p I primed T cells to preferred Derp I peptides analyzed by percent of response with an S.I. of at leasttwo within the individuals tested (above each bar), the mean T cellstimulation index (above each bar in parenthesis) and the ranked sum ofpeptide responses.

FIG. 16a is a graphic representation depicting the responses of T celllines from 9 patients primed in vitro to the Der f I protein andanalyzed for response to selected peptides of desired lengths derivedfrom the Der f I and Der p I protein allergens, by percent of responseswith an S.I. of at least 2 within the individuals tested and the mean Tcell stimulation index of responses with an S.I. of at least 2 for thepeptide.

FIG. 16b is a graphic representation derived from the same data shown inFIG. 16a showing the response of Der p I primed T cell lines topreferred Der f I and Der p I peptides by percent of responses with anS.I. of at least 2 within the individuals tested and the mean T cellstimulation index of responses with an S.I. of at least 2 for thepeptide.

FIG. 17a is a graphic representation depicting the response of T cellsfrom 29 patients primed in vitro to the Der p II protein and analyzedfor response to selected peptides of desired lengths derived from theDer p II protein, by the T cell stimulation index of a response with anS.I. of at least 2 for the peptide.

FIG. 17b is a graphic representation depicting the response of 30patients primed in vitro to the Der p II protein and analyzed forresponse to selected peptides derived from the Der p II protein bypercent of responses with an S.I. of at least 2 within the individualstested (above each bar), the mean T cell stimulation index (above eachbar in parenthesis) and the ranked sum of peptide response (X-axis).

FIG. 18a is a graphic representation depicting the response of T cellsfrom 10 patients primed in vitro to the Der f II protein and analyzedfor response to selected peptides of desired lengths derived from theDer p II and Der f II protein, by the T cell stimulation index of aresponse with an S.I. of at least 2 for the peptide.

FIG. 18b is a graphic representation derived from the same data shown inFIG. 18a showing the response of Der f II primed T cell lines topreferred Der p II peptides analyzed by percent responses with an S.I.of at least 2 within the individuals tested (above each bar), the mean Tcell stimulation index (above each bar in parenthesis) and the rankedsum of peptide responses (X-axis).

FIG. 18c is a graphic representation depicting the responses of T celllines from 4 matched patients primed in vitro with mite group I allergenand analyzed for response to preferred Der p I and Der f I peptides bypercent of responses with an S.I. of at least 2 within the individualstested and the mean T cell stimulation index of positive responses forthe peptide.

FIG. 18d is a graphic representation depicting the responses of T celllines from 6 matched patients primed in vitro with mite group IIallergen and analyzed for response to preferred Der p II peptides bypercent of responses with an S.I. of at least 2 within the individualstested and the mean stimulation index of positive responses for thepeptide.

FIG. 19a-19b are graphic representations of the results of a directbinding assay of IgE to affinity purified and recombinant Der p I andDer p II proteins and certain Der p I and Der p II overlapping peptides.

FIG. 20a-20b are graphic representations of the results of a directbinding assay of IgE to affinity purified and recombinant Der f I andDer f II proteins and certain Der f I and Der f II overlapping peptides.

FIG. 21a-21h are graphic representations of the results of a directbinding assay of IgE to a mixture of biochemically purified miteallergens (PMA) and various peptides derived from Der p I, Der f I, Derp II and Der f II.

FIG. 22 is a composite alignment of the amino acid sequences of five Derp I clones (a)-(e) which illustrates polymorphism in the Der p Iprotein. The numbering refers to the sequence of the Der p I(a) clone.The symbol (-) is used to indicate that the amino acid residue of a Derp I clone is identical to the corresponding amino acid sequences of Derp I(a) at that position. The amino acid sequences of these clonesindicate that there may be significant variation in Der p I, with fivepolymorphic amino acid residues found in the five sequences.

FIG. 23 is a composite alignment of the amino acid sequences of threeDer p II clones (c), (1) and (2) which illustrates polymorphism in theDer p II protein. The numbering refers to the sequence of the Der pII(c) clone. The symbol (.) is used to indicate that the amino acidresidue of a Der p II clone is identical to the corresponding amino acidresidue of Der p II (c) at that position.

FIG. 24 is a composite alignment of the amino acid sequences of six Derf II clones (i.e., pFL1 (SEQ ID NO: 157), pFL2 (SEQ ID NO: 158), MT3(SEQ ID NO: 159), MT5 (SEQ ID NO: 160), MT18 (SEQ ID NO: 161) and MT16(SEQ ID NO: 162)) which illustrates polymorphism is the Der f IIprotein. The numbering refers to the sequences of the Der f pLF1 clone.The symbol (.) is used to indicate that the amino acid residue of a Derf II clone is identical to the corresponding amino acid residue of Der fII pFL1 at that position.

FIG. 25 shows the nucleotide and amino acid sequences of a selectedpeptide which comprises various regions derived from Der p I, Der p IIand Der f I protein allergens.

FIG. 26 shows the nucleotide and amino acid sequences of a selectedpeptide which comprises various regions derived from Der p I, Der p IIand Der f I protein allergens.

FIG. 27 shows the nucleotide and amino acid sequences of a selectedpeptide which comprises various regions derived from Der p I, Der p IIand Der f I protein allergens.

FIG. 28 shows the amino acid sequences of modified peptides inaccordance with the invention.

FIG. 29 shows the amino acid sequences of various modified peptides ofthe invention.

FIG. 30 shows the amino acid sequences of various modified peptides ofthe invention.

FIG. 31a and 31b are graphic representations depicting the responses ofT cell lines from 18 patients (some matched) primed in vitro with mitegroup I allergen (Der p I) and analyzed for response to various peptidesand modified peptides of the invention, the y axis indicates thepositivity index for each peptide tested which is the mean S.I.multiplied by the percent of individuals tested responding to thepeptide with a mean S.I. of at least 2.

FIG. 32a and 32b are graphic representations depicting the responses ofT cell lines from 21 patients (some matched) primed in vitro with mitegroup II allergen (Der p II)and analyzed for response to variouspeptides and modified peptides of the invention, the y axis indicatesthe positivity index for each peptide tested which is the means S.I.multiplied by the by the percent of individuals tested responding to thepeptide with a mean S.I. of at least 2.

FIG. 33 shows the amino acid sequences of the "unique" peptides inaccordance with the invention including the novel "unique" peptides ofthe invention.

FIG. 34 shows overlapping peptides derived from Der p I and Der p IIprotein allergens used in T cell studies described herein.

FIG. 35 is a graphic representation depicting T cell responses to theoverlapping Der p I peptides shown in FIG. 2 and the Group I "unique"peptides, DPI-21.2 (SEQ. ID. NO. 28), DFI-22.2 (SEQ. ID. NO. 93),DFI-23.31(SEQ. ID. NO. 165), DFI-26.6 (SEQ. ID. NO. 168), shown in FIG.33. The mean S.I. shown above each bar (in parenthesis) as well as thepercentage of responses, the positivity index (mean S.I. multiplied bypercentage of responses), is the Y axis.

FIG. 36 is a graphic representation depicting T cell responses to theoverlapping Der p II peptides, DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14(SEQ. ID. NO. 180), and DPII-25.15 (SEQ. ID. NO. 188), shown in FIG. 2and the Der p II "unique" peptides shown in FIG. 2. The mean S.I. shownabove each bar (in parenthesis) as well as the percentage of responses,the positivity index (mean S.I. multiplied by percentage of responses),is the Y axis.

FIG. 37 is a pH-solubility profile of "unique" candidate peptidesDPI-21.2 (SEQ. ID. NO. 28), DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ.ID. NO. 165), DFI-26.6 (SEQ. ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169),DPII-22.14 (SEQ. ID. NO. 180) and DPII-25.15 (SEQ. ID. NO. 188), in 50mM sodium phosphate buffers with 5% mannitol. Solubility is measured inmg/ml (y axis) over a pH range of pH 5.5 to pH 8.5 at about 22° C.±2.

FIG. 38 is a pH-stability profile of candidate peptides DPI-21.2 (SEQ.ID. NO. 28), DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165),DFI-26.6 (SEQ. ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14(SEQ. ID. NO. 180) and DPII-25.15 (SEQ. ID. NO. 188) in an equalconcentration combination. Degradation of peptide is calculated as %degradation (determined by peak area using HPLC analysis) of peptideobserved after 24 hours at about 22° C.±2 and about 5° C., at varioustheoretical concentrations of 3.0, 2.0 and 1.0 mg/ml of peptide over apH range of 6.0 to 8.5.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides isolated peptides derived from the majorprotein allergens of the genus Dermatophagoides. As used herein, a"peptide" refers to an amino acid sequence having fewer amino acidresidues than the entire amino acid sequence of the protein from whichthe peptide was derived. The term "peptide" also refers to anyfunctional equivalents of a peptide or to any fragments or portions of apeptide. Peptides of the invention include peptides derived from Der p I(SEQ ID NO: 1 and 2), Der p II (SEQ ID NO: 3 and 4), Der f I (SEQ ID NO:5 and 6) and Der f II (SEQ ID NO: 7 and 8) which are antigenic (i.e.have the ability to induce an immune response). Preferably, peptides ofthe invention comprise at least one T cell epitope of the allergen.

Peptides comprising at least two regions, each region comprising atleast one T cell epitope of a protein allergen of the genusDermatophagoides are also within the scope of the invention. Each regionof such peptides is derived from the same or from different miteallergens. Isolated peptides or regions of isolated peptides, eachcomprising at least two T cell epitopes of a mite protein allergen arealso desirable for therapeutic effectiveness. Peptides which areimmunologically related (e.g., by antibody or T cell cross-reactivity)to peptides of the present invention are also within the scope of theinvention. Peptides immunologically related by antibody cross-reactivityare bound by antibodies specific for a peptide of a protein allergen ofthe genus Dermatophagoides. Peptides immunologically related by T cellcross-reactivity are capable of reacting with the same T cells as apeptide of the invention.

The present invention also provides modified peptides. As used herein,the term "modified peptide" refers to a peptide derived from a proteinallergen of the genus Dermatophagoides in which the amino acid sequencehas been altered by amino acid substitution, deletion or addition.Modified peptides also include those peptides which are modifications orvariations of peptides of the invention defined herein such as by aminoacid substitution, deletion or addition.

Isolated peptides of the invention can be produced by recombinant DNAtechniques in a host cell transformed with a nucleic acid having asequence encoding such peptide. The isolated peptides of the inventioncan also be produced by chemical synthesis. In certain situations,isolated peptides can be produced by chemical cleavage of the proteinallergen.

When a peptide is produced by recombinant techniques, host cellstransformed with a nucleic acid of the invention (or the functionalequivalent of the nucleic acid having a sequence encoding the peptide(or functional equivalent of the peptide) are cultured in a mediumsuitable for the cells. Peptides can be purified from cell culturemedium, host cells, or both using techniques known in the art forpurifying peptides and proteins including ion-exchange chromatography,gel filtration chromatography, ultrafiltration, electrophoresis orimmunopurification with antibodies specific for the peptide, the proteinallergen of the genus Dermatophagoides from which the peptide isderived, or a portion thereof. Isolated peptides of the invention aresubstantially free of cellular material or culture medium when producedby recombinant DNA techniques, or substantially free of chemicalprecursors or other chemicals when synthesized chemically or free ofother materials and reagents when produced by chemical cleavage.

The term "nucleic acid" as used herein is intended to include fragmentsor equivalents of nucleic acids of the invention. Nucleic acid sequencesused in any embodiment of this invention can be cDNA as describedherein, or alternatively, can be any oligodeoxynucleotide sequencehaving all or a portion of a sequence represented herein, or theirfunctional equivalents. Such oligodeoxynucleotide sequences can beproduced chemically or mechanically, using known techniques. Afunctional equivalent of an oligonucleotide sequence is one which is 1)a sequence capable of hybridizing to a complementary oligonucleotide towhich the sequence (or corresponding sequence portions) of SEQ ID NO: 1,SEQ ID NO: 3, SEQ ID NO: 5 and SEQ ID NO: 7, or fragments thereofhybridizes, or 2) the sequence (or corresponding sequence portion)complementary to SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5 and SEQ ID NO:7 and/or 3 a sequence which encodes a product (e.g., a polypeptide orpeptide) having the same functional characteristics of the productencoded by the sequence (or corresponding sequence portion) of SEQ IDNO: 1, SEQ ID NO: 3, SEQ ID NO: 5 and SEQ ID NO: 7. Whether a functionalequivalent must meet one or more criteria will depend on its use (e.g.,if it is to be used only as an oligoprobe, it need meet only the firstor second criteria and if it is to be used to produce a peptide of thepresent invention, it need only meet the third criterion).

As used herein, the "functional equivalent" of a peptide includespeptides having the same or enhanced ability to bind MHC, peptidescapable of stimulating the same T cell subpopulations, peptides havingthe same or increased ability to induce T cell responses such asstimulation (proliferation or cytokine secretion), peptides having thesame or increased ability to induce T cell non-responsiveness or reducedresponsiveness to a mite allergen, peptides having at least the samelevel of reduced IgE binding, and peptides which elicit at least thesame minimal level of IgE synthesis stimulating activity. Minimal IgEstimulating activity refers to IgE synthesis stimulating activity thatis less than the amount of IgE production elicited by a purified nativemite allergen

Preferred nucleic acids encode a peptide having at least about 50%homology to a peptide of the invention, more preferably at least about60% homology and most preferably at least about 70% homology with apeptide of the invention. Nucleic acids which encode peptides having atleast about 90%, more preferably at least about 95%, and most preferablyat least about 98-99% homology with a peptide of the invention are alsowithin the scope of the invention. Homology refers to sequencesimilarity between two peptides or between two nucleic acid molecules.Homology can be determined by comparing a position in each sequencewhich may be aligned for purposes of comparison. When a position in thecompared sequence is occupied by the same base or amino acid, thenmolecules are homologous at that position. A degree of homology betweensequences is a function of the number of matching or homologouspositions shared by the sequences.

The present invention provides expression vectors and host cellstransformed to express the nucleic acid sequences of the invention. Anucleic acid sequence coding for a peptide of the invention or at leastone fragment or portion thereof may be expressed in bacterial cells suchas E. coli, insect cells (baculovirus), yeast, or mammalian cells suchas Chinese hamster ovary cells (CHO). Suitable expression vectors,promoters, enhancers, and other expression control elements may be foundin Sambrook et al. Molecular Cloning: A Laboratory Manual, secondedition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.(1989). Other suitable expression vectors, promoters, enhancers, andother expression elements are known to those skilled in the art.Expression in mammalian, yeast or insect cells leads to partial orcomplete glycosylation of the recombinant material and formation of anyinter- or intra-chain disulfide bonds. Suitable vectors for expressionin yeast include YepSec 1 (Baldari et al. (1987) Embo J., 6: 229-234);pMFa (Kurjan and Herskowitz (1982) Cell, 30: 933-943); JRY88 (Schultz etal. (1987) Gene, 54: 113-123) and pYES2 (Invitrogen Corporation, SanDiego, Calif.). These vectors are freely available. Baculovirus andmammalian expression systems are also available. For example, abaculovirus system is commercially available (PharMingen, San Diego,Calif.) for expression in insect cells while the pMSG vector iscommerically available (Pharmacia, Piscataway, N.J.) for expression inmammalian cells.

For expression in E. coli, suitable expression vectors include, amongothers, pTRC (Amann et al. (1988) Gene, 69: 301-315); pGEX (Amrad Corp.,Melbourne, Australia); pMAL (N.E. Biolabs, Beverly, Mass.); pRIT5(Pharmacia, Piscataway, N.J.); pET-11 d (Novagen, Madison, Wis.) Jameelet al., (1990) J. Virol., 64:3963-3966; and pSEM (Knapp et al. (1990)BioTechniques, 8: 280-281). The use of pTRC, and pET-11 d, for example,will lead to the expression of unfused protein. The use of pMAL, pRIT5pSEM and pGEX will lead to the expression of allergen fused to maltose Ebinding protein (pMAL), protein A (pRIT5), truncated β-galactosidase(PSEM), or glutathione S-transferase (pGEX). When a mite peptide of theinvention is expressed as a fusion protein, it is particularlyadvantageous to introduce an enzymatic cleavage site at the fusionjunction between the carrier protein and a mite peptide. A mite peptideof the invention may then be recovered from the fusion protein throughenzymatic cleavage at the enzymatic site and biochemical purificationusing conventional techniques for purification of proteins and peptides.Suitable enzymatic cleavage sites include those for blood clottingFactor Xa or thrombin for which the appropriate enzymes and protocolsfor cleavage are commercially available from, for example, SigmaChemical Company, St. Louis, Mo. and N.E. Biolabs, Beverly, Mass. Thedifferent vectors also have different promoter regions allowingconstitutive or inducible expression with, for example, IPTG induction(PRTC, Amann et al., (1988) supra; pET-11 d, Novagen, Madison, Wis.) ortemperature induction (pRIT5, Pharmacia, Piscataway, N.J.). It may alsobe appropriate to express recombinant peptides of the invention indifferent E. coli hosts that have an altered capacity to degraderecombinantly expressed proteins (e.g. U.S. Pat. No. 4,758,512).Alternatively, it may be advantageous to alter the nucleic acid sequenceto use codons preferentially utilized by E. coli, where such nucleicacid alteration would not affect the amino acid sequence of theexpressed protein.

Host cells can be transformed to express the nucleic acid sequences ofthe invention using conventional techniques such as calcium phosphate orcalcium chloride co-precipitation, DEAE-dextran-mediated transfection,or electroporation. Suitable methods for transforming the host cells maybe found in Sambrook et al. supra, and other laboratory textbooks.

Inducible non-fusion expression vectors include pTrc (Amann et al.,(1988) Gene, 69:301-315) and pET11d (Studier et al., Gene ExpressionTechnology: Methods in Enzymology, 185, Academic Press, San Diego,Calif. (1990) 60-89). While target gene expression relies on host RNApolymerase transcription from the hybrid trp-lac fusion promoter inpTrc, expression of target genes inserted into pET11d relies ontranscription from the T7 gn10-lac 0 fusion promoter mediated bycoexpressed viral RNA polymerase (T7 gn1). This viral polymerase issupplied by host strains BL21(DE3) or HMS174(DE3) from a resident λprophage harboring a T7 gn1 under the transcriptional control of thelacUV 5 promoter.

One strategy to maximize recombinant expression of peptides of theinvention in E. coli is to express the protein in a host bacteria withan impaired capacity to proteolytically cleave the recombinant peptide(Gottesman, S., Gene Expression Technology: Methods in Enzymology, 185,Academic Press, San Diego, Calif. (1990) 119-128). Another strategywould be to alter the nucleic acid sequence of the desired peptide to beinserted into an expression vector so that the individual codons foreach amino acid would be those preferentially utilized in highlyexpressed E. coli proteins (Wada et al., (1992) Nuc. Acids Res.20:2111-2118). Such alteration of nucleic acid sequences of theinvention could be carried out by standard DNA synthesis techniques.

The nucleic acids of the invention can also be chemically synthesizedusing standard techniques. Various methods of chemically synthesizingpolydeoxynucleotides are known, including solid-phase synthesis which,like peptide synthesis, has been fully automated in commerciallyavailable DNA synthesizers (See e.g., Itakura et al. U.S. Pat. No.4,598,049; Caruthers et al. U.S. Pat. No. 4,458,066; and Itakura U.S.Pat. Nos. 4,401,796 and 4,373,071, incorporated by reference herein).

To obtain isolated peptides of the present invention, a mite allergen isdivided into non-overlapping peptides of desired length or overlappingpeptides of desired lengths as discussed in Example III which can beproduced recombinantly, synthetically, or in certain situations, bychemical cleavage of the allergen. Peptides comprising at least one Tcell epitope are capable of eliciting a T cell response, such asstimulation (i.e. proliferation or lymphokine secretion) and/or arecapable of inducing T cell non-responsiveness. To determine peptidescomprising at least one T cell epitope, isolated peptides are tested by,for example, T cell biology techniques, to determine whether thepeptides elicit a T cell response or induce T cell non-responsivenss.Those peptides found to elicit a T cell response or induce T cellnon-responsiveness are defined as having T cell stimulating activity.

Screening peptides of the invention for human T cell stimulatingacitivity can be accomplished using one or more of several differentassays. For example, in vitro, T cell stimulatory activity is assayed bycontacting a peptide of the invention with an antigen presenting cellwhich presents appropriate MHC molecules in a T cell culture.Presentation of a peptide of the invention in association withappropriate MHC molecules to T cells, in conjunction with the necessarycostimulation has the effect of transmitting a signal to the T cell thatinduces the production of increased levels of cytokines, particularly ofinterleukin-2 and interleukin-4. The culture supernatant can be obtainedand assayed for interleukin-2 or other known cytokines. For example, anyone of several conventional assays for interleukin-2 can be employed,such as the assay described in Proc. Natl. Acad. Sci USA, 86:1333 (1989)the pertinent portions of which are incorporated herein by reference. Akit for an assay for the production of interferon is also available fromGenzyme Corporation (Cambridge, Mass.).

A common assay for T cell proliferation entails measuring tritiatedthymidine incorporation. The proliferation of T cells can be measured invitro by determining the amount of ³ H-labeled thymidine incorporatedinto the replicating DNA of cultured cells. Therefore, the rate of DNAsynthesis and, in turn, the rate of cell division can be quantified.

A peptide may also be screened for the ability to reduce T cellresponsiveness. The ability of a peptide known to stimulate T cells, toinhibit or completely block the activity of a purified nativeDermatophagoides protein allergen or portion thereof and induce a stateof T cell nonresponsiveness or reduced T cell responsiveness, can bedetermined using subsequent attempts at stimulation of the T cells withantigen presenting cells that present a native Dermatophagoides allergenfollowing exposure to a peptide of the invention. If the T cells areunresponsive to the subsequent activation attempts, as determined byinterleukin-2 synthesis and T cell proliferation, a state ofnonresponsiveness has been induced. See, e.g., Gimmi, et al. (1993)Proc. Natl. Acad. Sci USA, 90:6586-6590; and Schwartz (1990) Science,248:1349-1356, for assay systems that can be used as the basis for anassay in accordance with the present invention.

Additionally, peptides comprising "cryptic epitopes" may be determinedand are also within the scope of this invention. Cryptic epitopes arethose determinants in a protein antigen which, due to processing andpresentation of the native protein antigen to the appropriate MHCmolecule, are not normally revealed to the immune system. However, apeptide comprising a cryptic epitope is capable of causing T cells tobecome non-responsive, and when a subject is primed with the peptide, Tcells obtained from the subject will proliferate in vitro in response tothe peptide or the protein antigen from which the peptide is derived.Peptides which comprise at least one cryptic epitope derived from aprotein antigen are referred to herein as "cryptic peptides". To confirmthe presence of cryptic epitopes in the above-described T cellproliferation assay, antigen-primed T cells are cultured in vitro in thepresence of each peptide separately to establish peptide-reactive T celllines. A peptide is considered to comprise at least one cryptic epitopeif a T cell line can be established with a given peptide and T cells arecapable of proliferation upon challenge with the peptide and the proteinantigen from which the peptide is derived.

As discussed above and in the Examples, human T cell stimulatingactivity can be tested by culturing T cells obtained from an individualsensitive to a mite allergen, (i.e., an individual who has an IgEmediated immune response to a mite allergen) with a peptide derived fromthe allergen and determining whether proliferation of T cells occurs inresponse to the peptide as measured, e.g., by cellular uptake oftritiated thymidine. Stimulation indices for responses by T cells topeptides can be calculated as the maximum CPM in response to a peptidedivided by the control CPM. A T cell stimulation index (S.I.) equal toor greater than two times the background level is considered "positive."Positive results are used to calculate the mean stimulation index foreach peptide for the group of peptides tested. Preferred peptides ofthis invention comprise at least one T cell epitope and have a mean Tcell stimulation index of greater than or equal to 2.0. A peptide havinga T cell stimulation index of greater than or equal to 2.0 is considereduseful as a therapeutic agent. Preferred peptides have a mean T cellstimulation index of at least 2.5, more preferably at least 3.5, evenmore preferably at least 4.0, and most preferably at least 5.0.

In addition, preferred peptides have a positivity index (P.I.) of atleast about 100, more preferably at least 150, even more preferably atleast about 200 and most preferably at least about 250. The positivityindex for a peptide is determined by multiplying the mean T cellstimulation index by the percent of individuals, in a population ofindividuals sensitive to house dust mite (e.g., preferably at least 9individuals, more preferably at least 16 individuals or more, morepreferably at least 29 individuals or more, or even more preferably atleast 30 individuals or more), who have T cells that respond to thepeptide. Thus, the positivity index represents both the strength of a Tcell response to a peptide (S.I.) and the frequency of a T cell responseto a peptide in a population of individuals sensitive to house dustmite. For example, as shown in FIG. 37, peptide DP I-1 has a mean S.I.of 4.7 and 73% of positive responses in the group of individuals testedresulting in a positivity index of 343.1. As shown in FIG. 37, peptidesof Der p I having a positivity index of at least about 150 and a mean Tcell stimulation index of at least about 4 include: DP I-1 (SEQ ID NO:9); DP I-21.1 (SEQ ID NO: 27); DP I-21.2 (SEQ ID NO: 28); DP I-23.1 (SEQID NO: 33); DP I-23.2 (SEQ ID NO: 34); DP I-25.2 (SEQ ID NO: 36); and DPI-26.1 (SEQ ID NO: 37).

In order to determine precise T cell epitopes by, for example, finemapping techniques, a peptide having T cell stimulating activity andthus comprising at least one T cell epitope as determined by T cellbiology techniques is modified by addition or deletion of amino acidresidues at either the amino or carboxy terminus of the peptide andtested to determine a change in T cell reactivity to the modifiedpeptide. If two or more peptides which share an area of overlap in thenative protein sequence are found to have human T cell stimulatingactivity, as determined by T cell biology techniques, additionalpeptides can be produced comprising all or a portion of such peptidesand these additional peptides can be tested by a similar procedure.Following this technique, peptides are selected and producedrecombinantly or synthetically or by chemical cleavage. Peptides areselected based on various factors, including the strength of the T cellresponse to the peptide (e.g., stimulation index), the frequency of theT cell response to the peptide in a population of individuals sensitiveto house dust mite, and the potential cross-reactivity of the peptidewith other allergens of the genus Dermatophagoides. The physical andchemical properties of these selected peptides (e.g., solubility,stability) are examined to determine whether the peptides are suitablefor use in therapeutic compositions or whether the peptides requiremodification as described herein. The ability of the selected peptidesor selected modified peptides to stimulate human T cells (e.g., induceproliferation, lymphokine secretion) is determined. Peptide DF I-22.2(FIG. 36) is one example of the product of this process. This peptide isderived from amino acid residues 40-64 of Der f I. This peptide wasconstructed from information obtained during T cell studies of theinitial overlapping peptides (Example III). These initial studiesindicated that overlapping peptides DP I-4 and DP I-11.1, which togethercovered amino acid residues 40-60 of Der p I (which is homologous to Derf I in that region), both gave a positive response in the T cell assays,indicating that at least one epitope existed in that region (see ExampleV). Thus, several peptides were constructed (for "fine mapping"purposes) which covered the general region of overlapping peptides DPI-4 and DP I-11.1, including DF I-22.2, DF I-22.4 and DP I-22.4 amongothers. FIG. 31a-b shows the relationship of the peptides under the sameassay conditions. FIG. 31b shows that overlapping peptides DP I-4 and DPI-11.1 give positive responses indicating the presence of T cellreactivity. FIG. 31a shows that peptides DF I-22.2, DP I-22.4 and DFI-22.4 (amino acid residues 36-64 of Der f I, and Der p I) which are allbased on the general region covered by overlapping peptides DP I-4 andDP I-11.1, also have positive responses indicating that each of thesepeptides also comprises at least one T cell epitope. In addition, it maybe desirable to further modify the "fine map" peptides such as DF I-22.2for purposes of increasing solubility or stability. For example, peptideDF I-22.2 which has the amino acid sequence VAATESAYLAYRNTSLDLSEQELVDmay be further modified by removing the first amino acid, valine (V),from the amino terminus and also removing one or both of the the lasttwo amino acid residues (valine (V) and aspartic acid (D) from thecarboxy terminus of the peptide. This modification may increase thestability or solubility of the peptide. In addition charged amino acidsmay replace the deleted amino acid residues to further increase thepeptide's solubility.

The results in FIGS. 31a-b and 32a-b show a number of other successfulexamples of the fine mapping techniques combined with furthermodifications of the fine map peptides. For example, as shown in FIG.31b, peptides DP I-1 and DP I-2 each give a positive response. Togetherthese peptides cover amino acid residues 1-29 of Der p I. Peptides DPI-21.1 and DP I-21.2 were constructed as fine map peptides to cover this1-29 region. Peptides DP I-21.1 and DP I-21.2 both give a positiveresponse indicating that at least one epitope from the overlappingregion covered by peptides DP I-1 and DP I-2 was preserved. Moreover,the results in FIG. 31a indicate that peptide DP I-21.2 has a higherpositivity index then either of the overlapping peptides, DP I-1 and DPI-2 (FIG. 31b), which makes it a very useful peptide. It may bedesirable to further modify DP I-21.2 to increase its solubility orstability for the purposes of including it in a therapeutic composition.Modified peptide DP I-21.7 is an example of such a modification in whichthe "core" of peptide DP I-21.2 remains essentially unchanged except acharged amino acid is added to the amino terminus and a methionineinternal to the peptide is replaced with a leucine resulting in peptideDP I-21.7. Similarly positive T cell studies with other fine mappeptides shown in FIGS. 31a and 32a (i.e. DP I-23.1, DP I-26.1 DPII-20.0, DP II-22, DP II-25) and modified for thereof (i.e. DP I-23.31,DP I-23.35, DP I-26.6, DP II-20.9, DP II-20.11, DP II-20.10, DPII-8, DPII-22.19, DP II-22.22, DP II-22.23, DP II-22.14, DP II-25.9) show thatregions of reactivity can be identifed with overlapping peptides (FIGS.31b and 32b), and fine map peptides may be constructed from thoseregions which preserve and perhaps enhance the T cell reactivitypreviously defined in the overlapping region, and using the amino acidsequences of the fine map peptides as a "core", further modificationsmay be made to the fine map peptides to impart other desiredcharacteristics to the peptide such as increased solubility andstability, which is important if the peptide is to be used as atherpeutic composition. Examples of specific modifications to peptidesfor purposes of increased solubility, stability etc. are discussedlater.

If a peptide of the invention is to be used for therapeutic purposes, itis preferred that the peptide does not bind immunoglobulin E (IgE) orbinds IgE to a substantially lesser extent (i.e. at least 100-fold lessbinding and more preferably, at least 1,000-fold less binding) than theprotein allergen from which the peptide is derived binds IgE. The majorcomplications of standard immunotherapy are IgE-mediated responses suchas anaphylaxis. Immunoglobulin E is a mediator of anaphylactic reactionswhich result from the binding and cross-linking of antigen to IgE onmast cells or basophils and the release of mediators (e.g., histamine,serotonin, eosinophil chemotacic factors) in allergic ("atopic")patients (see, Example XVI). Thus, anaphylaxis in a substantialpercentage of a population of individuals sensitive to house dust mitecould be avoided by the use in immunotherapy of a peptide or peptideswhich do not bind IgE in a substantial percentage (e.g., at least about75%) of a population of individuals sensitive to a house dust miteallergen, or if the peptide binds IgE, such binding does not result inthe release of mediators from mast cells or basophils. The risk ofanaphylaxis could be reduced by the use in immunotherapy of a peptide orpeptides which have reduced IgE binding. Moreover, peptides which haveminimal IgE stimulating activity are desirable for therapeuticeffectiveness. Minimal IgE stimulating activity refers to IgE productionthat is less than the amount of IgE production and/or IL-4 productionstimulated by the native protein allergen (e.g., Der p I).

If a peptide of the invention binds IgE, and is to be used as atherapeutic agent, it is preferable that such binding does not result inthe release of mediators (e.g. histamines) from mast cells or basophils.To determine whether a peptide which binds IgE results in the release ofmediators, a histamine release assay can be performed using standardreagents and protocols obtained for example, from Amac, Inc. (Westbrook,Me.). Briefly, a buffered solution of a peptide to be tested is combinedwith an equal volume of whole heparinized blood from an allergicsubject. After mixing and incubation, the cells are pelleted and thesupernatants are processed and analyzed using a radioimmunoassay todetermine the amount of histamine released.

If a peptide of the invention is to be used as a diagnostic reagent, itis not necessary that the peptide or protein have reduced IgE bindingactivity compared to the native Dermatophagoides allergen (e.g. Der p I,Der p II, Der f I or Der f II). IgE binding activity of peptides can bedetermined by, for example, an enzyme linked immunosorbent assay (ELISA)using, for example, sera obtained from a subject, (i.e., an allergicsubject) that has been previously exposed to the native Dermatophagoidesallergen. Briefly, a peptide to be tested is coated onto wells of amicrotiter plate. After washing and blocking the wells, antibodysolution consisting of the plasma of an allergic subject who has beenexposed to the peptide being tested or the protein from which it wasderived is incubated in the wells. The plasma is generally depleted ofIgG before incubation. A labeled secondary antibody is added to thewells and incubated. The amount of IgE binding is then quantified andcompared to the amount of IgE bound by a purified nativeDermatophagoides protein allergen. Alternatively, the binding activityof a peptide can be determined by Western blot analysis. For example, apeptide to be tested is run on a polyacrylamide gel using SDS-PAGE. Thepeptide is then transferred to nitrocellulose and subsequently incubatedwith sera from an allergic subject. After incubation with the labeledsecondary antibody, the amount of IgE bound is then determined andquantified.

Another assay which can be used to determine IgE binding activity of apeptide is a competition ELISA assay. Briefly, an IgE antibody pool isgenerated by combining plasma from house dust mite allergic subjectsthat have been shown by direct ELISA to have IgE reactive with nativeDermatophagoides protein allergen. This pool is used in ELISAcompetition assays to compare IgE binding to native Dermatophagoidesprotein allergen to the peptide tested. IgE binding for the nativeDermatophagoides protein allergen and the peptide being tested isdetermined and quantified.

A peptide of the invention, when administered to a house dustmite-sensitive individual in a therapeutic regimen, is capable ofmodifying the allergic response of the individual to the allergen.Particularly, peptides of the invention comprising at least one T cellepitope of a mite allergen or at least two regions derived from a miteallergen, each comprising at least one T cell epitope, when administeredto an individual sensitive to house dust mite are capable of modifyingthe T cell response, the B cell response or both the T cell and B cellresponse of the individual to the allergen. As used herein, modificationof the allergic response of an individual sensitive to a house dust miteallergen can be defined as non-responsiveness or diminution in symptomsupon exposure to a mite allergen, as determined by standard clinicalprocedures (see e.g., Varney et al., British Medical Journal,302:265-269 (1990)) including diminution in mite protein allergeninduced asthmatic symptoms. As referred to herein, a diminution insymptoms includes any reduction in allergic response of an individual tothe allergen after the individual has completed a treatment regimen witha peptide of the invention. This diminution may be subjective (i.e. thepatient feels more comfortable in the presence of the allergen).Diminution in symptoms can be determined clinically as well, usingstandard skin tests as is known in the art.

As a result of the work described herein, peptides derived from miteallergens which have T cell stimulating activity and thus comprise atleast one T cell epitope have been produced. T cell epitopes arebelieved to be involved in initiation and perpetuation of the immuneresponse to a protein allergen which is responsible for the clinicalsymptoms of allergy. These T cell epitopes are thought to trigger earlyevents at the level of the T helper cell by binding to an appropriateHLA molecule on the surface of an antigen presenting cell andstimulating the relevant T cell subpopulation. These events lead to Tcell proliferation, lymphokine secretion, local inflammatory reactions,recruitment of additional immune cells to the site, and activation ofthe B cell cascade leading to production of antibodies. One isotype ofthese antibodies, IgE, is fundamentally important to the development ofallergic symptoms and its production is influenced early in the cascadeof events, at the level of the T helper cell, by the nature of thelymphokines secreted. A T cell epitope is the basic element or smallestunit of recognition by a T cell receptor, where the epitope comprisesamino acids essential to receptor recognition. Amino acid sequenceswhich mimic those of the T cell epitopes and which modify the allergicresponse to protein allergens are also within the scope of thisinvention.

Exposure of house dust mite patients to isolated peptides of the presentinvention which comprise at least one T cell epitope, and are derivedfrom Dermatophatophagoides protein allergens, in a non-immunogenic form,may cause T cell non-responsiveness of appropriate T cell subpopulationssuch that they become unresponsive to the protein allergen and do notparticipate in stimulating an immune response upon such exposure or maycause reduced T cell responsiveness. In addition, administration of apeptide of the invention which comprises at least one T cell epitope maymodify the lymphokine secretion profile as compared with exposure to thenaturally-occurring protein allergen or portion thereof (e.g. result ina decrease of IL-4 and/or an increase in IL-2). Furthermore, exposure tosuch peptide may influence T cell subpopulations which normallyparticipate in the response to the allergen such that these T cells aredrawn away from the site(s) of normal exposure to the allergen (e.g.,nasal mucosa, skin, and lung) towards the site(s) of therapeuticadministration of the fragment or protein allergen. This redistributionof T cell subpopulations may ameliorate or reduce the ability of anindividual's immune system to stimulate the usual immune response at thesite of normal exposure to the allergen, resulting in a diminution inallergic symptoms.

The isolated peptides of the invention can be used in methods ofdiagnosing, treating and preventing allergic reactions to house dustmite allergen or a cross reactive protein allergen. Thus the presentinvention provides therapeutic compositions comprising peptides derivedfrom house dust mite allergens and a pharmaceutically acceptable carrieror diluent. Administration of the therapeutic compositions of thepresent invention to an individual to be desensitized can be carried outusing known techniques. Peptides of the invention may be administered toan individual in combination with, for example, an appropriate diluent,a carrier and/or an adjuvant or incomplete adjuvant. Pharmaceuticallyacceptable diluents include saline and aqueous buffer solutions.Pharmaceutically acceptable carriers include polyethylene glycol (Wie etal. (1981) Int. Arch. Allergy Appl. Immunol., 64:84-99) and liposomes(Strejan et al. (1984) J. Neuroimmunol., 7: 27).

The therapeutic compositions of the invention are administered to housedust mite allergen-sensitive individuals or individuals sensitive to anallergen which is immunologically cross-reactive with house dust miteallergen. For purposes of inducing T cell non responsiveness,therapeutic compositions of the invention are preferably administered innon-immunogenic form, e.g. which does not contain adjuvant. While notintending to be limited to any theory, it is believed that T cell nonresponsivness or reduced T cell responsiveness is induced as a result ofnot providing an appropriate costimulatory signal sometimes referred toas a "second signal" Briefly, it is believed that stimulation of T cellsrequires two types of signals, the first is the recognition by the Tcell via the T cell receptor of appropriate MHC-associated processedantigens on antigen presenting cells (APCs) and the second type ofsignal is referred to as a costimulatory signal(s) or "second signal"which may be provided by certain competent APCs. When a composition ofthe invention is administered without adjuvant, it is believed thatcompetent APCs which are capable of producing the second signal orcostimulatory signal are not engaged in the stimulation of appropriate Tcells therefore resulting in T cell nonresponsiveness or reduced T cellresponsiveness. In addition, there are a number of antibodies or otherreagents capable of blocking the delivery of costimulatory signals suchas the "second signal" which include, but are not limited to B7(including B7-1, B7-2, and BB-1), CD28, CTLA4, CD40 CD40L CD54 and CD11a/18 (Jenkins and Johnson, Current Opinion in Immunology, 5:361-367(1993), and Clark and Ledbetter, Nature, 367:425-428 (1994)) Thus, apeptide of the invention may be administered in nonimmunogenic form asdiscussed above, in conjunction with a reagent capable of blockingcostimulatory signals such that the level of T cell nonresponsiveness isenhanced.

Administration of the therapeutic compositions of the present inventionto an individual to be desensitized can be carried out using knownprocedures at dosages and for periods of time effective to reducesensitivity (i.e., reduce the allergic response) of the individual tothe allergen. Effective amounts of the therapeutic compositions willvary according to factors such as the degree of sensitivity of theindividual to Japanese cedar pollen, the age, sex, and weight of theindividual, and the ability of the protein or fragment thereof to elicitan antigenic response in the individual. The active compound (i.e.,protein or fragment thereof) may be administered in a convenient mannersuch as by injection (subcutaneous, intravenous, etc.), oraladministration, inhalation, transdermal application, or rectaladministration. Depending on the route of administration, the activecompound may be coated within a material to protect the compound fromthe action of enzymes, acids and other natural conditions which mayinactivate the compound.

For example, preferably about 1 μg- 3 mg and more preferably from about20-500 μg of active compound (i.e., protein or fragment thereof) perdosage unit may be administered by injection. Dosage regimen may beadjusted to provide the optimum therapeutic response. For example,several divided doses may be administered daily or the dose may beproportionally reduced as indicated by the exigencies of the therapeuticsituation.

To administer protein or peptide by other than parenteraladministration, it may be necessary to coat the protein with, orco-administer the protein with, a material to prevent its inactivation.For example, protein or fragment thereof may be administered in anincomplete adjuvant, co-administered with enzyme inhibitors or inliposomes. Enzyme inhibitors include pancreatic trypsin inhibitor,diisopropylfluorophosphate (DEP) and trasylol. Liposomes includewater-in-oil-in-water CGF emulsions as well as conventional liposomes(Strejan et al., (1984) J. Neuroimmunol., 7:27).

The active compound may also be administered parenterally orintraperitoneally. Dispersions can also be prepared in glycerol, liquidpolyethyline glycols, and mixtures thereof and in oils. Under ordinaryconditions of storage and use, these preparations may contain apreservative to prevent the growth of microorganisms.

Pharmaceutical compositions suitable for injectable use include sterileaqueous solutions (where water soluble) or dispersions and sterilepowders for the extemporaneous preparation of sterile injectablesolutions of dispersion. In all cases, the composition must be sterileand must be fluid to the extent that easy syringability exists. It mustbe stable under the conditions of manufacture and storage and must bepreserved against the contaminating action of microorganisms such asbacteria and fungi. The carrier can be a solvent or dispersion mediumcontaining, for example, water, ethanol, polyol (for example, glyceral,propylene glycol, and liquid polyetheylene glycol, and the like),suitable mixtures thereof, and vegetable oils. The proper fluidity canbe maintained, for example, by the use of a coating such as licithin, bythe maintenance of the required particle size in the case of dispersionand by the use of surfactants. Prevention of the action ofmicroorganisms can be achieved by various antibacterial and antifungalagents, for example, parabens, chlorobutanol, phenol, ascorbic acid,thirmerosal, and the like. In many cases, it will be preferable toinclude isotonic agents, for example, sugars, polyalcohols such asmanitol and sorbitol or sodium chloride in the composition. Prolongedabsorption of the injectable compositions can be brought about,including in the composition, an agent which delays absorption, forexample, aluminum monostearate and gelatin.

Sterile injectable solutions can be prepared by incorporating activecompound (i.e., protein or peptide) in the required amount in anappropriate solvent with one or a combination of ingredients enumeratedabove, as required, followed by filtered sterilization. Generally,dispersions are prepared by incorporating the active compound into asterile vehicle which contains a basic dispersion medium and therequired other ingredients from those enumerated above. In the case ofsterile powders for the preparation of sterile indectable solutions, thepreferred methods of preparation are vacuum drying and freeze-dryingwhich yields a powder of the active ingredient (i.e., protein orpeptide) plus any additional desired ingredient from a previouslysterile-filtered solution thereof.

When a peptide is suitably protected, as described above, the peptidemay be orally administered, for example, with an inert diluent or anassimilable edible carrier. The peptide and other ingredients may alsobe enclosed in a hard or soft shell gelatin capsule, compressed intotablets, or incorporated directly into the individual's diet. For oraltherapeutic administration, the active compound may be incorporated withexcipients and used in the form of ingestible tablets, buccal tablets,troches, capsules, elixirs, suspensions, syrups, wafers, and the like.Such compositions and preparations should contain at least 1% by weightof active compound. The percentage of the composition and preparationsmay, of course, be varied and may conveniently be between about 5 to 80%of the weight of the unit. The amount of active compound in suchtherapeutically useful compositions is such that a suitable dosage willbe obtained. Preferred compositions or preparations according to thepresent invention are prepared so that an oral dosage unit containsbetween from about 10 μg to about 200 mg of active compound.

The tablets, troches, pills, capsules and the like may also contain thefollowing: a binder such as gum gragacanth, acacia, corn starch orgelatin; excipients such as dicalcium phosphate; a disintegrating agentsuch as corn starch, potato starch, alginic acid and the like; alubricant such as magnesium stearate; and a sweetening agent such assucrose, lactose or saccharin or a flavoring agent such as peppermint,oil of wintergreen, or cherry flavoring. When the dosage unit form is acapsule, it may contain, in addition to materials of the above type, aliquid carrier. Various other materials may be present as coatings or tootherwise modify the physical form of the dosage unit. For instance,tablets, pills, or capsules may be coated with shellac, sugar or both. Asyrup or elixir may contain the active compound, sucrose as a sweeteningagent, methyl and propylparabens as preservative, a dye and flavoringsuch as cherry or orange flavor. Of course, any material used inpreparing any dosage unit form should be pharmaceutically pure andsubstantially non-toxic in the amounts employed. In addition, the activecompound may be incorporated into sustained-release preparations andformulations.

As used herein "pharmaceutically acceptable carrier" includes any andall solvents, dispersion media, coatings, antibacterial and antifungalagents, isotonic and absorption delaying agents, and the like. The useof such media and agents for pharmaceutically active substances is wellknown in the art. Except insofar as any conventional media or agent isincompatible with the active compound, use thereof in the therapeuticcompositions is contemplated. Supplementary active compounds can also beincorporated into the compositions.

It is especially advantageous to formulate parenteral compositions indosage unit form for ease of administration and uniformity of dosage.Dosage unit from as used herein refers to physically discrete unitssuited as unitary dosages for the mammalian subjects to be treated; eachunit containing a predetermined quantity of active compound calculatedto produce the desired therapeutic effect in association with therequired pharmaceutical carrier. The specification for the novel dosageunit forms of the invention are dictated by and directly dependent on(a) the unique characteristics of the active compound and the particulartherapeutic effect to be achieved, and (b) the limitations inherent inthe art of compounding such an active compound for the treatment ofsensitivity in individuals.

The invention further includes isolated allergenic proteins or fragmentsthereof that are immunologically related to a Dermatophagoides proteinallergen or peptide thereof, such as by antibody cross-reactivitywherein the isolated allergenic proteins or fragments thereof arecapable of binding to antibodies specific for the peptides of theinvention, or by T cell cross-reactivity wherein the isolated allergenicproteins or fragments thereof are capable of stimulating T cellsspecific for the peptides of this invention.

Peptides encoded by the cDNA of the present invention can be used, forexample as "purified" allergens. Such purified allergens are useful inthe standardization of allergen extracts which are currently keyreagents for the clinical diagnosis and treatment of sensitivity tohouse dust mite allergens.

Another aspect of the invention pertains to an antibody specificallyreactive with a peptide of the invention. The antibodies of thisinvention can be used to standardize allergen extracts or to isolate thenaturally-occurring or native forms of protein allergens derived fromthe genus Dermatophagoides or other immunologically related allergens.

The term antibody as used herein is intended to include fragmentsthereof which are also specifically reactive with a peptide of theinvention. Antibodies can be fragmented using conventional techniquesand the fragments screened for utility in the same manner as describedabove for whole antibodies. For example, F(ab')₂ fragments can begenerated by treating antibody with pepsin. The resulting F(ab')₂fragment can be treated to reduce disulfide bridges to produce Fab'fragments. The antibody of the present invention is further intended toinclude bispecific and chimeric molecules having an anti-Der p I, Der pII, Der f I or Der f II portion.

Another aspect of this invention provides isolated T cell clones andisolated soluble T cell receptors specifically reactive with peptides ofthe invention. Monoclonal T cell populations (i.e., T cells geneticallyidentical to one another and expressing identical T cell receptors) canbe derived from an individual sensitive to house dust mite proteinallergen, followed by repetitive in vitro stimulation with house dustmite protein allergen in the presence of MHC-matched antigen-presentingcells. Single house dust mite protein allergen MHC responsive cells canthen be cloned by limiting dilution and permanent lines expanded andmaintained by periodic in vitro restimulation. Alternatively, house dustmite protein allergen specific T--T hybridomas can be produced by atechnique similar to B cell hybridoma production. For example, a mammal,such as a mouse can be immunized with a peptide of the invention, Tcells from the mammal can be purified and fused with an autonomouslygrowing T cell tumor line. From the resulting hybridomas, cellsresponding to a peptide of the invention are selected and cloned.Procedures for propagating monoclonal T cell populations are describedin Cellular and Molecular Immunology (Abul K. Abbas et al. ed.), W. B.Saunders Company, Philadelphia, Pa. (1991) page 139. Soluble T cellreceptors specifically reactive with a peptide of the invention orfragments thereof can be obtained by immunoprecipitation using anantibody against the T cell receptor as described in Immunology: ASynthesis (Second Edition), Edward S. Golub et al., ed., SinauerAssociates, Inc., Sunderland, Mass. (1991) pages 366-269.

T cell clones specifically reactive with a peptide of the invention canbe used to isolate and molecularly clone the gene encoding the relevantT cell receptor. In addition, a soluble T cell receptor specificallyreactive with a peptide of the invention can be used to interfere withor inhibit antigen-dependent activation of the relevant T cellsubpopulation, for example, by administration to an individual sensitiveto house dust mite protein allergen. Antibodies specifically reactivewith such a T cell receptor can be produced according to knowntechniques. Such antibodies can be used to block or interfere with the Tcell interaction with peptides presented by MHC.

Through use of the peptides of the present invention, preparations ofconsistent, well-defined composition and biological activity can be madeand administered for therapeutic purposes (e.g. to modify the allergicresponse of a house dust mite sensitive individual). Administration ofsuch peptides or protein may, for example, modify B-cell response to ahouse dust mite protein allergen, T-cell response to a house dust miteprotein allergen, or both responses. Isolated peptides can also be usedto study the mechanism of immunotherapy of house dust mite proteinallergy and to design modified derivatives or analogues useful inimmunotherapy.

Work by others has shown that high doses of allergens generally producethe best results (i.e., best symptom relief). However, many people areunable to tolerate large doses of allergens because of allergicreactions to the allergens. A peptide can be designed in such a mannerto have the same or enhanced therapeutic properties as the correspondingnaturally-occurring allergen but have reduced side effects (especiallyanaphylactic reactions) can be produced. These can be, for example, apeptide of the present invention.

It is also possible to modify the structure of a peptide of theinvention for such purposes as increasing solubility, enhancingtherapeutic or preventive efficacy, or stability (e.g., shelf life exvivo, and resistance to proteolytic degradation in vivo). A modifiedpeptide can be produced in which the amino acid sequence has beenaltered, such as by amino acid substitution, deletion, or addition, tomodify immunogenicity and/or reduce allergenicity, or to which acomponent has been added for the same purpose.

For example, a peptide can be modified so that it maintains the abilityto induce T cell anergy and bind MHC proteins without the ability toinduce a strong proliferative response or possibly, any proliferativeresponse when administered in immunogenic form. In this instance,critical binding residues for the T cell receptor can be determinedusing known techniques (e.g., substitution of each residue anddetermination of the presence or absence of T cell reactivity). Thoseresidues shown to be essential to interact with the T cell receptor canbe modified by replacing the essential amino acid with another,preferably similar amino acid residue (a conservative substitution)whose presence is shown to enhance, diminish but not eliminate, or noteffect T cell reactivity. In addition, those amino acid residues whichare not essential for T cell receptor interaction can be modified bybeing replaced by another amino acid whose incorporation may enhance,diminish or not effect T cell reactivity but does not eliminate bindingto relevant MHC.

Additionally, peptides of the invention can be modified by replacing anamino acid shown to be essential to interact with the MHC proteincomplex with another, preferably similar amino acid residue(conservative substitution) whose presence is shown to enhance, diminishbut not eliminate or not effect T cell activity. In addition, amino acidresidues which are not essential for interaction with the MHC proteincomplex but which still bind the MHC protein complex can be modified bybeing replaced by another amino acid whose incorporation may enhance,not effect, or diminish but not eliminate T cell reactivity. Preferredamino acid substitutions for non-essential amino acids include, but arenot limited to substitutions with alanine, glutamic acid, or a methylamino acid.

Another example of a modification of peptides is substitution ofcysteine residues preferably with serine, threonine, leucine or glutamicacid to minimize dimerization via disulfide linkages. For example, thestability of a peptide of Der p II or Der f II which includes the firstten amino acid residues of either allergen can be enhanced by replacingthe cysteine located at the 8th amino acid residue, preferably withalanine, or glutamic acid, or alternatively with serine or threonine.

In order to enhance stability and/or reactivity, peptides can also bemodified to incorporate one or more polymorphisms in the amino acidsequence of a protein allergen resulting from natural allelic variationas shown in FIGS. 25-27. Additionally, D-amino acids, non-natural aminoacids or non-amino acid analogues can be substituted or added to producea modified peptide within the scope of this invention. Furthermore,peptides of the present invention can be modified using the polyethyleneglycol (PEG) method of A. Sehon and co-workers (Wie et al. supra) toproduce a peptide conjugated with PEG. In addition, PEG can be addedduring chemical synthesis of a peptide of the invention. Modificationsof peptides or portions thereof can also include reduction/alkylation(Tarr in: Methods of Protein Microcharacterization, J. E. Silver ed.Humana Press, Clifton, N.J., pp 155-194 (1986)); acylation (Tarr,supra); esterification (Tarr, supra); chemical coupling to anappropriate carrier (Mishell and Shiigi, eds, Selected Methods inCellular Immunology, WH Freeman, San Francisco, Calif. (1980); U.S. Pat.No. 4,939,239); or mild formalin treatment (Marsh International Archivesof Allergy and Applied Immunology, 41:199-215 (1971)).

In another embodiment, peptides within an allergen group can be modifiedto enhance T cell reactivity. Given the cross-reactivity within theGroup I and Group II allergens, a peptide of one group allergen whichmay be less reactive than a peptide of another group allergencorresponding in amino acid position can have one or more amino acidssubstituted with one or more amino acids from the corresponding peptide(e.g., peptide DF II-1, residue seventeen methionine! in the Der f IIamino acid sequence can be substituted with the amino acid located atresidue seventeen in Der p II leucine! to enhance the reactivity of theDer f II peptide). Additionally, peptides can be modified to incorporatea polymorphism in the amino acid sequence of a protein allergenresulting from natural allelic variation. Modification of peptides toinclude one or more of these polymorphisms may result in enhancedstability and/or reactivity.

To facilitate purification and potentially increase solubility ofpeptides of the invention, it is possible to add reporter group(s) tothe peptide backbone. For example, poly-histidine can be added to apeptide to purify the peptide on immobilized metal ion affinitychromatography (Hochuli, E. et al., Bio/Technology, 6:1321-1325 (1988)).In addition, specific endoprotease cleavage sites can be introduced, ifdesired, between a reporter group and amino acid sequences of a peptideto facilitate isolation of peptides free of irrelevant sequences.

In order to successfully desensitize an individual to a peptide, it maybe necessary to increase the solubility of a peptide for use in bufferedaqueous solutions, such as pharmaceutically acceptable carriers ordiluents, by adding functional groups to the peptide, terminal portionsof the peptide, or by not including hydrophobic T cell epitopes orregions containing hydrophobic epitopes in the peptides or hydrophobicregions of the protein or peptide. For example, to increase solubility,charged amino acids or charged amino acid pairs or triplets may be addedto the carboxy or amino terminus of the peptide. Examples of chargedamino acids include, arginine (R), lysine (K), histidine (H), glutamicacid (E), and aspartic acid (D).

To potentially aid proper antigen processing of T cell epitopes within apeptide, canonical protease sensitive sites can be recombinantly orsynthetically engineered between regions, each comprising at least one Tcell epitope. For example, charged amino acid pairs, such as KK or RR,can be introduced between regions within a peptide during recombinantconstruction of the peptide. The resulting peptide can be renderedsensitive to cathepsin and/or other trypsin-like enzymes cleavage togenerate portions of the peptide containing one or more T cell epitopes.In addition, such charged amino acid residues can result in an increasein solubility of a peptide.

Site-directed mutagenesis of DNA encoding a peptide of the invention canbe used to modify the structure of the peptide. Such methods may involvePCR (Ho et al., Gene, 77:51-59 (1989)) or total synthesis of mutatedgenes (Hostomsky, Z., et al., Biochem. Biophys. Res. Comm.,161:1056-1063 (1989)). To enhance bacterial expression, theaforementioned methods can be used in conjunction with other proceduresto change the eucaryotic codons in DNA constructs encoding peptides ofthe invention to ones preferentially used in E. coli.

Specific examples of peptides of the invention modified in accordancewith one or more of the modification procedures discussed above includebut are not limited to modifications to peptide DPI-23. 1 as shown inFIG. 28 and to peptide DPII-22 as shown in FIG. 28. More specifically,modifications to peptides 23.1 include the addition of charged aminoacids (23.2) charged amino acid pairs (23.1) to increase solubility andreplacement of a cysteine residue with serine 23.1.3 or glutamic acid(23.1.4) to increase solubility. Changes to peptide 22 include theaddition of charged amino acid pairs (22.1 and 22.2) to increasesolubility.

It is now also possible to design an agent or a drug capable of blockingor inhibiting the ability of house dust mite allergen to induce anallergic reaction in house dust mite allergen-sensitive individuals.Such agents could be designed, for example, in such a manner that theywould bind to relevant anti-Der p or Der f protein allergen IgEs, thuspreventing IgE-allergen binding and subsequent mast cell degranulation.Alternatively, such agents could bind to cellular components of theimmune system, resulting in suppression or desensitization of theallergic response to house dust mite allergens.

Peptides of the present invention can also be used for detecting anddiagnosing sensitivity to house dust mite protein allergens. Forexample, this could be done by combining blood or blood productsobtained from an individual to be assessed for sensitivity to house dustmite protein allergens with an isolated antigenic peptide or peptides ofthe invention, under conditions appropriate for binding of components inthe blood (e.g., antibodies, T cells, B cells) with the peptide(s) anddetermining the extent to which such binding occurs. Other diagnosticmethods for allergic diseases which the peptides of the presentinvention can be used include radio-allergergosorbent test (RAST), paperradioimmunosorbent test (PRIST), enzyme linked immunosorbent assay(ELISA), radioimmunoassays (RIA), immuno-radiometric assays (IRMA),luminescence immunoassays (LIA), histamine release assays and IgEimmunoblots.

The presence in individuals of IgE specific for at least one proteinallergen of the genus Dermatophagoides and the ability of T cells of theindividuals to respond to T cell epitope(s) of the protein allergen canbe determined by administering to the individuals an Immediate TypeHypersensitivity test and a Delayed Type Hypersensitiity test. Theindividuals are administered an Immediate Type Hypersensitivity test(see e.g. Immunology (1985) Roitt, I. M., Brostoff, J., Male, D. K.(eds), C. V. Mosby Co., Gower Medical Publishing, London, N.Y., pp.19.2-19.18; pp. 22.1-22.10) utilizing a peptide of the protein allergen,or a modified form of the peptide, each of which binds IgE specific forthe allergen. The same individuals are administered a Delayed TypeHypersensitivity test prior to, simultaneously with, or subsequent toadministration of the Immediate Type Hypersensitivity test. Of course,if the Immediate Type Hypersensitivity test is administered prior to theDelayed Type Hypersensitivity test, the Delayed Type Hypersensitivitytest would be given to those individuals exhibiting a specific ImmediateType Hypersensitivity reaction. The Delayed Type Hypersensitivity testutilizes a modified form of the protein allergen or a portion thereof,the protein allergen produced recombinantly, or peptide derived from theprotein allergen, each of which has human T cell stimulating activityand each of which does not bind IgE specific for the allergen in asubstantial percentage of the population of individuals sensitive to theallergen (e.g., at least about 75%). Those individuals found to haveboth a specific Immediate Type Hypersensitivity reaction and a specificDelayed Type Hypersensitivity reaction are diagnosed as havingsensitivity to house dust mite protein allergen and may, if need be,administered a therapeutically effective amount of a therapeuticcomposition of the invention. The therapeutic composition may comprise apeptide of the invention and a pharmaceutically acceptable carrier ordiluent.

For therapeutic purposes, isolated peptides of the invention comprise atleast one T cell epitope of a protein allergen of the genusDermatophagoides and accordingly, the peptide comprises at leastapproximately seven amino acid residues of the protein allergen.Therapeutic compositions of the invention may comprise at least two Tcell epitopes of a mite allergen. Accordingly, isolated peptides of theinvention comprising at least two T cell epitopes comprise at leastapproximately eight amino acid residues, and preferably fifteen aminoacid residues. Additionally, therapeutic compositions of the inventionpreferably comprise a sufficient percentage of the T cell epitopes ofthe entire protein allergen (i.e.at least about 40% and more preferablyabout 60% of the T cell reactivity to Der p and/or Der f allergens) areincluded in the composition such that a therapeutic regimen ofadministration of the composition to an individual sensitive to housedust mite, results in T cells of the individual being renderednonresponsive to the protein allergen or protein allergens.Synthetically produced peptides of the invention comprising up toapproximately forty-five amino acid residues in length, and mostpreferably up to approximately thirty amino acid residues in length areparticularly desirable as increases in length may result in difficultyin peptide synthesis as well as retention of an undesirable property(e.g., immunoglobulin binding or enzymatic activity) due to maintenanceof conformational similarity between the peptide and the proteinallergen from which it is derived

Preferred peptides comprise all or a portion of the areas of major Tcell reactivity within the Der p I, Der f I, Der p II and Der f IIprotein allergens, i.e., Region 1, Region 2, Region 3, Region 4, Region5, Region 6a, Region 6b, Region 7, Region 8, Region 9 and Region 10.Each area is defined as follows: Region 1 comprises amino acid residues1-28 of the Der p I and Der f I protein allergens; Region 2 comprisesamino acid residues 36-68 of the Der p I and Der f I protein allergens;Region 3 comprises amino acid residues 74-109 of the Der p I and Der f Iprotein allergens; Region 4 comprises amino acid residues 118-139 of theDer p I and Der f I protein allergens; Region 5 comprises amino acidresidues 141-166 of the Der p I and Der f I protein allergens; Region 6acomprises amino acid residues 161-185 of the Der p I and Der f I proteinallergens; Region 6b comprises amino acid 173-201 of the Der p I and Derf I allergens; Region 7 comprises amino acid residues 1-26 of the Der pII and Der f II protein allergens; Region 8 comprises amino acidresidues 33-67 of the Der p II and Der f II protein allergens; Region 9comprises amino acid residues 79-104 of the Der p II and Der f IIprotein allergens and Region 10 comprises amino acid residues 107-129 ofthe Der p II and Der f II protein allergens.

Preferred peptides derived from the Der p I protein comprise all or aportion of the following peptides: DP I-1 (SEQ ID NO: 9); DP I-2 (SEQ IDNO: 10); DP I-3 (SEQ ID NO: 11); DP I-4 (SEQ ID NO: 12); DP I-11.1 (SEQID NO: 13); DP I-12.1 (SEQ ID NO: 14); DP I-5 (SEQ ID NO: 15); DP I-13(SEQ ID NO: 17); DP I-14 (SEQ ID NO: 18); DP I-15 (SEQ ID NO: 19); DPI-6.1 (SEQ ID NO: 20); DP I-7.1 (SEQ ID NO: 21); DP I-8 (SEQ ID NO: 22);DP I-9 (SEQ ID NO: 23); DP I-16 (SEQ ID NO: 24); DP I-10 (SEQ ID NO:25); DP I-17 (SEQ ID NO: 26); DP I-21.1 (SEQ ID NO: 27); DP I-21.2 (SEQID NO: 28); DP I-22.1 (SEQ ID NO: 29); DP I-22.2 (SEQ ID NO: 30); DPI-22.3 (SEQ ID NO: 31); DP I-22.4 (SEQ ID NO: 32); DP I-23.1 (SEQ ID NO:33); DP I-23.2 (SEQ ID NO: 34); DP I-25.1 (SEQ ID NO: 35); DP I-25.2(SEQ ID NO: 36); DP I-26.1 (SEQ ID NO: 37); DP I-27.1 (SEQ ID NO: 38);DP I-28.1 (SEQ ID NO: 39); and DP I-28.2 (SEQ ID NO: 40), wherein theportion of the peptide has a mean T cell stimulation index substantiallyequivalent to, or greater than the mean T cell stimulation index of thepeptide from which it is derived as shown in FIG. 37 and FIG. 13. Morepreferably, peptides derived from the Der p I protein comprise all or aportion of the following peptides: DP I-21.2, DP I-22.2, DPI-23.1, DPI-25.2, DPI-26.1, DPI-27.1 and DP I-28.1, and most preferably, peptidesderived from the Der p I protein comprises all or a portion of DPI-21.2, DP I-23.1 and DP I-26.1, wherein the portion of the peptide hasa mean T cell stimulation index substantially equivalent to, or greaterthan the mean T cell stimulation index of the peptide from which it isderived as shown in FIG. 37 and FIG. 13.

Preferred peptides derived from the Der f I, the Der p II and the Der fII proteins include: DF I-1 (SEQ ID NO: 72); DF I-2.1 (SEQ ID NO: 73);DF I-3 (SEQ ID NO: 74); DF I-4 (SEQ ID NO: 75); DF I-11 (SEQ ID NO: 76);DF I-12 (SEQ ID NO: 77); DF I-5 (SEQ ID NO: 78); DF I-13 (SEQ ID NO:79); DF I-14 (SEQ ID NO: 80); DF I-15 (SEQ ID NO: 81); DF I-6 (SEQ IDNO: 82); DF I-7 (SEQ ID NO: 83); DF I-8.1 (SEQ ID NO: 84); DF I-8 (SEQID NO: 85); DF I-9 (SEQ ID NO: 86); DF I-16 (SEQ ID NO: 87); DF I-10(SEQ ID NO: 88); DF I-17 (SEQ ID NO: 89); DF I-21.1 (SEQ ID NO: 90); DFI-21.2 (SEQ ID NO: 91); DF I-22.1 (SEQ ID NO: 92); DF I-22.2 (SEQ ID NO:93); DF I-22.4 (SEQ ID NO: 94); DF I-23.1 (SEQ ID NO: 95); DF I-23.2(SEQ ID NO: 96); DF I-25.1 (SEQ ID NO: 97); DF I-25.2 (SEQ ID NO: 98);DF I-26.1 (SEQ ID NO: 99); DF I-27.1 (SEQ ID NO: 100); DF I-28.1 (SEQ IDNO: 101); DF I-28.2 (SEQ ID NO: 102); DP II-20 (SEQ ID NO: 50); DPII-20.1 (SEQ ID NO: 51); DP II-20.2 (SEQ ID NO: 52); DP II-20.3 (SEQ IDNO: 53); DP II-20.4 (SEQ ID NO: 54); DP II-20.5 (SEQ ID NO: 55); DP II20.6 (SEQ ID NO: 56); DP II-1 (SEQ ID NO: 41); DP II-2 (SEQ ID NO: 42);DP II-3.1 (SEQ ID NO: 43); DP II-4 (SEQ ID NO: 44); DP II-5 (SEQ ID NO:45); DP II-6 (SEQ ID NO: 46); DP II-7 (SEQ ID NO: 47); DP II-8 (SEQ IDNO: 48); DP II-9 (SEQ ID NO: 49); DP II-1.1 (SEQ ID NO: 57); DP II-1.2(SEQ ID NO: 58); DP II-2.1 (SEQ ID NO: 59); DP II-2.2 (SEQ ID NO: 60);DP II-2.3 (SEQ ID NO: 61); DP II-21 (SEQ ID NO: 62); DP II-22 (SEQ IDNO: 63); DP II-26 (SEQ ID NO: 64); DP II-26.1 (SEQ ID NO: 65); DP II-23(SEQ ID NO: 66); DP II-23.1 (SEQ ID NO: 67); DP II-24 (SEQ ID NO: 68);DP II-25 (SEQ ID NO: 69); DP II-25.1 (SEQ ID NO: 70); DP II-25.2 (SEQ IDNO: 71); DF II-1 (SEQ ID NO: 103); DF II-2 (SEQ ID NO: 104); DF II-13.1(SEQ ID NO: 105); DF II-3.1 (SEQ ID NO: 106); DF II-4.5 (SEQ ID NO:107); DF II-4.3 (SEQ ID NO: 108); DF II-15 (SEQ ID NO: 109); DF II-16(SEQ ID NO: 110); DF II-17 (SEQ ID NO: 111); DF II-18 (SEQ ID NO: 112);DF II-19 (SEQ ID NO: 113); DF II-19.1 (SEQ ID NO: 114); DF II-21 (SEQ IDNO: 115); and DF II-22 (SEQ ID NO: 116), or portions of the peptidescomprising at least one T cell epitope. Preferably, a portion of apeptide derived from Der f I, Der p II and Der f II has a mean T cellstimulation index equivalent to or greater than the mean T cellstimulation index of the peptide from which it is derived as shown inFIG. 10, FIG. 15a, FIG. 15b and FIG. 16. More preferably, peptidesderived from the Der p II and Der f I proteins comprise all or a portionof the following peptides: DF I-22.2 (ID SEQ NO: 93), DP II-20.6 (ID SEQNO: 56), DP II-20.0 (ID SEQ NO: 50), DP II-22 (ID SEQ NO: 63), DP II-24(ID SEQ NO: 68) and DP II-25.2 (ID SEQ NO: 71).

Preferred modified peptides derived from Der p I, Der p II, and Der f IIprotein allergens include: DP I-21.7 (SEQ ID NO: 120), DP I-23.10 (SEQID NO: 121), DP I-23.11 (SEQ ID NO: 124), DP I-23.12 (SEQ ID NO: 125),DP I-23.5 (SEQ ID NO: 126), DP I-23.6 (SEQ ID NO: 127), DP I-23.7 (SEQID NO: 128), DP I-23.8 (SEQ ID NO: 129), DP I-23.9 (SEQ ID NO: 130), DPI-26.2 (SEQ ID NO: 134), DP II-20.7 (SEQ ID NO: 138), DP II-22.6 (SEQ IDNO: 139), DP II-22.3 (SEQ ID NO: 140), DP II-22.4 (SEQ ID NO: 141), DPI-22.5 (SEQ ID NO: 142), DP II-25.3 (SEQ ID NO: 148), DP II-25.4 (SEQ IDNO: 149), DP I-23.13 (SEQ ID NO: 122), DP I-23.14 (SEQ ID NO: 123), DPI-23.15 (SEQ ID NO: 131), DP I-23.16 (SEQ ID NO: 132), DP I-23.17 (SEQID NO: 133), DP I-26.3 (SEQ ID NO: 135), DP I-26.4 (SEQ ID NO: 136), DPI-26.5 (SEQ ID NO: 137), DP II-22.7 (SEQ ID NO: 143), DP II-22.8 (SEQ IDNO: 144), DP II-22.9 (SEQ ID NO: 145), DP II-22.10 (SEQ ID NO: 146), DPII-22.11 (SEQ ID NO: 147), DP I-23.32 (SEQ ID NO: 163), DPI-23.33 (SEQID NO: 164) DP I-23.31 (SEQ ID NO: 165), DP I-23.34 (SEQ ID NO: 166), DPI-23.35 (SEQ ID NO: 167), DP I-26.6 (SEQ ID NO: 168), DP II-20.9 (SEQ IDNO: 169), DP II-20.11 (SEQ ID NO: 169), DP II-20.10 (SEQ ID NO: 170) DPII-20.8 (SEQ ID NO: 171), D,P II-22.19 (SEQ ID NO: 172), DP II-22.20(SEQ ID NO: 173), DP II-22.21 (SEQ ID NO: 174), DP II-22.22 (SEQ ID NO:175), DP II-22.26 (SEQ ID NO: 176), DP II-22.23 (SEQ ID NO: 177), DPII-22.24 (SEQ ID NO: 178), DP II-22.25 (SEQ ID NO: 179), DP II-22.14(SEQ ID NO: 180), DF II-25.11 (SEQ ID NO: 182), DP II-25.9 (SEQ ID NO:183), DF II-25.10 (SEQ ID NO: 184), DF II-25.13 (SEQ ID NO: 186), DPII-25.14 (SEQ ID NO: 187), DP II-25.15 (SEQ ID NO: 188), DP II-25.16(SEQ ID NO: 189), DP II-25.17 (SEQ ID NO: 190), DP II-25.18 (SEQ ID NO:191) all as shown in FIGS. 29 and 30.

Preferably modified peptides derived from Der p I, Der p II, and Der fII protein allergens include: DP I-21.7 (SEQ ID NO: 120), DP I-23.31(SEQ ID NO: 165), DP I-23.35 (SEQ ID NO: 167), DP I-26.6 (SEQ ID NO:168), DP II-20.9 (SEQ ID NO: 169), DP II 20.11 (SEQ ID NO: 169), DPII-20.10 (SEQ ID NO: 170), DP II-8 (SEQ ID NO: 171), DP II-22.19 (SEQ IDNO: 172), DP II-22.22 (SEQ ID NO: 175), DP II-22.23 (SEQ ID NO: 177), DPII-22.14 (SEQ ID NO: 180), DP II-25.9 (SEQ ID NO: 183), DP II-25.14 (SEQID NO: 187), DP II-25.15 (SEQ ID NO: 188), DP II-25.16 (SEQ ID NO: 189),DP II-25.17 (SEQ ID NO: 190), and DP II-25.18 (SEQ ID NO: 191).

One embodiment of the present invention features a peptide of a proteinallergen of the genus Dermatophagoides. The peptide or portion thereofcomprises at least one T cell epitope of the protein allergen and has aformula X_(n) --Y--Z_(m). According to the formula, Y is an amino acidsequence selected from the group consisting of: DF II-21.1 (SEQ ID NO:90); DF I-21.2 (SEQ ID NO: 91); DF I-22.1 (SEQ ID NO: 92); DF I-22.2(SEQ ID NO: 93); DF I-22.4 (SEQ ID NO: 94); DF I-23.1 (SEQ ID NO: 95);DF I-23.2 (SEQ ID NO: 96); DF I-25.1 (SEQ ID NO: 97); DF I-25.2 (SEQ IDNO: 98); DF I-26.1 (SEQ ID NO: 99); DF I-27.1 (SEQ ID NO: 100); DFI-28.1 (SEQ ID NO: 101); DF I-28.2 (SEQ ID NO: 102); DF I-1 (SEQ ID NO:72); DP II-20 (SEQ ID NO: 50); DP II-20.1 (SEQ ID NO: 51); DP II-20.2(SEQ ID NO: 52); DP II-20.3 (SEQ ID NO: 53); DP II-20.4 (SEQ ID NO: 54);DP II-20.5 (SEQ ID NO: 55); DP II 20.6 (SEQ ID NO: 56); DP II-1 (SEQ IDNO: 41); DP II-1.1 (SEQ ID NO: 57); DP II-1.2 (SEQ ID NO: 58); DP II-2.1(SEQ ID NO: 59); DP II-2.2 (SEQ ID NO: 60); DP II-2.3 (SEQ ID NO: 61);DP II-21 (SEQ ID NO: 62); DP II-22 (SEQ ID NO: 63); DP II-26 (SEQ ID NO:64); DP II-26.1 (SEQ ID NO:65); DP II-23 (SEQ ID NO: 66); DP II-23.1(SEQ ID NO: 67); DP II-24 (SEQ ID NO: 68); DP II-25 (SEQ ID NO: 69); DPII-25.1 (SEQ ID NO: 70); DP II-25.2 (SEQ ID NO: 71); DF II-1 (SEQ ID NO:103); DF II-2 (SEQ ID NO: 104); DF II-13.1 (SEQ ID NO: 105); DF II-3.1(SEQ ID NO: 106); DF II-4.5 (SEQ ID NO: 107); DF II-4.3 (SEQ ID NO:108); DF II-15 (SEQ ID NO: 109); DF II-16 (SEQ ID NO: 110); DF II-17(SEQ ID NO: 111); DF II-18 (SEQ ID NO: 112); DF II-19 (SEQ ID NO: 113);DF II-19.1 (SEQ ID NO: 114); DF II-21 (SEQ ID NO: 115); and DF II-22(SEQ ID NO: 116). In addition, X_(n) are amino acid residues contiguousto the amino terminus of Y in the amino acid sequence of the proteinallergen and Z_(m) are amino acid residues contiguous to the carboxyterminus of Y in the amino acid sequence of the protein allergen.Preferably, the amino acids comprising the amino terminus of X and thecarboxy terminus of Z are selected from charged amino acids, i.e.,arginine (R), lysine (K), histidine (H), glutamic acid (E) or asparticacid (D); amino acids with reactive side chains, e.g., cysteine (C),asparagine (N) or glutamine (Q); or amino acids with sterically smallside chains, e.g., alanine (A) or glycine (G). In the formula, n ispreferably 0-30 and m is preferably 0-30. Preferably n and m are 0-5,and most preferably n+m is less than 10. Preferably, the peptide orportion thereof has a mean T cell stimulation index equivalent to orgreater than the mean T cell stimulation index of Y (see Examples andFigs).

In the above equation, Y may also be selected from the group of modifiedpeptides consisting of: DP I-21.7 (SEQ ID NO: 120); DP I-23.10 (SEQ IDNO: 121); DP I-23.11 (SEQ ID NO: 124); DP I-23.12 (SEQ ID NO: 125); DPI-23.5 (SEQ ID NO: 126); DP I-23.6 (SEQ ID NO: 127); DP I-23.7 (SEQ IDNO: 128); DP I-23.8 (SEQ ID NO: 129); DP I-23.9 (SEQ ID NO: 130); DPI-26.2 (SEQ ID NO: 134); DP II-20.7 (SEQ ID NO: 138); DP II-22.6 (SEQ IDNO: 139); DP II-22.3 (SEQ ID NO: 140); DP II-22.4 (SEQ ID NO: 141); DPII-22.5 (SEQ ID NO: 142); DP II-25.3 (SEQ ID NO: 148); DP II-25.4 (SEQID NO: 149); DP I-23.13 (SEQ ID NO: 122); DP I-23.14 (SEQ ID NO: 123);DP I-23.15 (SEQ ID NO: 131); DP I-23.16 (SEQ ID NO: 132); DP I-23.17(SEQ ID NO: 133); DP I-26.3 (SEQ ID NO: 135); DP I-26.4 (SEQ ID NO:136); DP I-26.5 (SEQ ID NO: 137); DP II-22.7 (SEQ ID NO: 143); DPII-22.8 (SEQ ID NO: 144); DP II-22.9 (SEQ ID NO: 145); DP II-22.10 (SEQID NO: 146); DP II-22.11 (SEQ ID NO: 147); DP I-23.32 (SEQ ID NO: 163),DPI-23.33 (SEQ ID NO: 164), DP I-23.31 (SEQ ID NO: 165), DP I-23.34 (SEQID NO: 166), DP I-23.35 (SEQ ID NO: 167), DP I-26.6 (SEQ ID NO: 168), DPII-20.9 (SEQ ID NO: 169), DP II-20.11 (SEQ ID NO: 169), DP II-20.10 (SEQID NO: 170), DP II-20.8 (SEQ ID NO: 171), D,P II-22.19 (SEQ ID NO: 172),DP II-22.20 (SEQ ID NO: 173), DP II-22.21 (SEQ ID NO: 174), DP II-22.22(SEQ ID NO: 175), DP II-22.26 (SEQ ID NO: 176), DP II-22.23 (SEQ ID NO:177), DP II-22.24 (SEQ ID NO: 178), DP II-22.25 (SEQ ID NO: 179), DPII-22.14 (SEQ ID NO: 180), DF II-25.11 (SEQ ID NO: 182), DP II-25.9 (SEQID NO: 183), DF II-25.10 (SEQ ID NO: 184), DF II-25.13 (SEQ ID NO: 186),DP II-25.14 (SEQ ID NO: 187), DP II-25.15 (SEQ ID NO: 188), DP II-25.16(SEQ ID NO: 189), DP II-25.17 (SEQ ID NO: 190), DP II-25.18 (SEQ ID NO:191) all as shown in FIGS. 29 and 30.

Another embodiment of the present invention provides peptides comprisingat least two regions, each region comprising at least one T cell epitopeof a protein allergen of the genus Dermatophagoides and accordingly,each region comprises at least approximately seven amino acid residues).These peptides comprising at least two regions can comprise as manyamino acid residues as desired and preferably comprise at least about14, even more preferably about 30, and most preferably at least about 40amino acid residues of a mite allergen. Each region of such peptidepreferably comprises up to 45 amino acid residues in length, morepreferably up to 40 residues in length and most preferably up to 30amino acid residues in length as increases in length of a region mayresult in difficulty in peptide synthesis as well as retention of anundesirable property (e.g., immunoglobulin binding or enzymaticactivity) due to maintenance of conformational similarity between thepeptide and the protein allergen from which it is derived. If desired,the amino acid sequences of the regions can be produced and joined by alinker to increase sensitivity to processing by antigen-presentingcells. Such linker can be any non-epitope amino acid sequence or otherappropriate linking or joining agent. To obtain preferred peptidescomprising at least two regions, each comprising at least one T cellepitope, the regions are arranged in a configuration different from anaturally-occurring configuration of the regions in the allergen or acombination of different mite protein allergens. For example, theregions containing T cell epitope(s) can be arranged in a noncontiguousconfiguration and can preferably be derived from the same proteinallergen or a combination of protein allergens. Noncontiguous is definedas an arrangement of regions containing T cell epitope(s) which isdifferent than that of an amino acid sequence present in the proteinallergen from which the regions are derived. Furthermore, thenoncontiguous regions containing T cell epitopes can be arranged in anonsequential order (e.g., in an order different from the order of theamino acids of the native protein allergen from which the regioncontaining T cell epitope(s) are derived in which amino acids arearranged from an amino terminus to a carboxy terminus). A peptide cancomprise at least 15%, at least 30%, at least 50% or up to 100% of the Tcell epitopes of a mite allergen but does not comprise the whole proteinsequence of the allergen.

The individual peptide regions can be produced and tested to determinewhich regions bind immunoglobulin E specific for a mite allergen andwhich of such regions would cause the release of mediators (e.g.,histamine) from mast cells or basophils. Those peptide regions found tobind immunoglobulin E and cause the release of mediators from mast cellsor basophils in greater than approximately 10-15% of the allergic seratested are preferably not included in the peptide regions arranged toform peptides of the invention.

Preferred peptides of the invention comprise two or more regions derivedfrom the same or from different mite allergens (e.g. Der p I, Der p II,Der f I and Der f II). For example, one region can be derived from Der pI and one region can be derived from Der p II; one region can be derivedfrom Der p I and one region can be derived from Der f I; one region canbe derived from Der p II and one region can be derived from Der f I; oneregion can be derived from Der p II and one region can be derived fromDer f II; one region can be derived from Der p I and one region can bederived from Der f II; and one region can be derived from Der f I andone region can be derived from Der f II. In addition, the regions can bederived from the same protein allergen, e.g., Der p I and Der p I, etc.

Regions of a peptide of the invention preferably comprise all or aportion of the above discussed preferred areas of major T cellreactivity within each mite allergen (i.e., Regions 1-6a-6b of Der p Iand Der f I and Regions 7-10 of Der p II and Der f II). For example, oneregion can comprise all or a portion of Region I (amino acid residues1-28 of Der p I or Der f I) and one region can comprise all or a portionof Region 2 (amino acid residues 36-68 of Der p I or Der f I). Peptidesof the invention can comprise all or a portion of two or more of theseRegions (i.e., Regions 1-10) and preferred resulting peptides do notbind IgE and cause the release of mediators from mast cells orbasophils. Preferred peptides derived from Der p I and Der f I compriseall or a portion of Region 1, Region 2, Region 3 and optionally Region4. Preferred peptides derived from Der p II and Der f II comprise all ora portion of Region 7 and Region 8 and, Region 10. Further, if one ofthese Regions is found to bind IgE and cause the release of mediatorsfrom mast cells or basophils, then it is preferred that the peptide notcomprise such Regions but rather comprises various Regions derived fromsuch Regions which do not bind IgE or cause release of mediators frommast cells or basophils.

Examples of preferred regions include all or a portion of the followingamino acid sequences: DP I-21.1 (SEQ ID NO: 27); DP I-21.2 (SEQ ID NO:28); DP I-22.1 (SEQ ID NO: 29); DP I-22.2 (SEQ ID NO: 30); DP I-22.3(SEQ ID NO: 31); DP I-22.4 (SEQ ID NO: 32); DP I-23.1 (SEQ ID NO: 33);DP I-23.2 (SEQ ID NO: 34); DP I-25.1 (SEQ ID NO: 35); DP I-25.2 (SEQ IDNO: 36); DP I-26.1 (SEQ ID NO: 37); DP I-27.1 (SEQ ID NO: 38); DP I-28.1(SEQ ID NO: 39); DP I-28.2 (SEQ ID NO: 40); DP I-1 (SEQ ID NO: 9); DFI-1 (SEQ ID NO: 72); DF I-21.1 (SEQ ID NO: 90); DF I-21.2 (SEQ ID NO:91); DF I-22.1 (SEQ ID NO: 92); DF I-22.2 (SEQ ID NO: 93); DF I-22.4(SEQ ID NO: 94); DF I-23.1 (SEQ ID NO: 95); DF I-23.2 (SEQ ID NO: 96);DF I-25.1 (SEQ ID NO: 97); DF I-25.2 (SEQ ID NO: 98); DF I-26.1 (SEQ IDNO: 99); DF I-27.1 (SEQ ID NO: 100); DF I-28.1 (SEQ ID NO: 101); DFI-28.2 (SEQ ID NO: 102); DP II-20 (SEQ ID NO: 50); DP II-20.1 (SEQ IDNO: 51); DP II-20.2 (SEQ ID NO: 52); DP II-20.3 (SEQ ID NO: 53); DPII-20.4 (SEQ ID NO: 54); DP II-20.5 (SEQ ID NO: 55); DP II-20.6 (SEQ IDNO: 56); DP II-1 (SEQ ID NO: 41); DP II-1.1 (SEQ ID NO: 57); DP II-1.2(SEQ ID NO: 58); DP II-2.1 (SEQ ID NO: 59); DP II-2.2 (SEQ ID NO: 60);DP II-2.3 (SEQ ID NO: 61); DP II-21 (SEQ ID NO: 62); DP II-22 (SEQ IDNO: 63); DP II-26 (SEQ ID NO: 64); DP II-26.1 (SEQ ID NO: 65); DP II-23(SEQ ID NO: 66); DP II-23.1 (SEQ ID NO: 67); DP II-24 (SEQ ID NO: 68);DP II-25 (SEQ ID NO: 69); DP II-25.1 (SEQ ID NO: 70); DP II-25.2 (SEQ IDNO: 71); DF II-1 (SEQ ID NO: 103) DF II-2 (SEQ ID NO: 104); DF II-13.1(SEQ ID NO: 105); DF II-3.1 (SEQ ID NO: 106); DF II-4.5 (SEQ ID NO:107); DF II-4.3 (SEQ ID NO: 108); DF II-15 (SEQ ID NO: 109); DF II-16(SEQ ID NO: 110); DF II-17 (SEQ ID NO: 111); DF II-18 (SEQ ID NO: 112);DF II-19 (SEQ ID NO: 113); DF II-19.1 (SEQ ID NO: 114); DF II-21 (SEQ IDNO: 115); and DF II-22 (SEQ ID NO: 116), the amino acid sequences ofsuch regions being shown in FIG. 3 and FIG. 36, or portions of saidregions comprising at least one T cell epitope.

Preferred peptides comprise various combinations of two or more regions,each region comprising all or a portion of the above-discussed preferredareas of major T cell reactivity. Preferred peptides comprise acombination of two or more regions (each region having an amino acidsequence as shown in FIG. 3 and FIG. 36) including: DP I-22.1 (SEQ IDNO: 29) and DP I-25.1 (SEQ ID NO: 35); DP I-21.1 (SEQ ID NO: 27) and DPI-25.2 (SEQ ID NO: 36); DP I-22.1 (SEQ ID NO: 29) and DP I-1 (SEQ ID NO:9); DP I-21.1 (SEQ ID NO: 27), DP I-22.1 (SEQ ID NO: 29), and DP I-25.2(SEQ ID NO: 36); DP I-21.2 (SEQ ID NO: 28), DP I-22.1 (SEQ ID NO: 29),and DP I-23.1 (SEQ ID NO: 39); DP I-1 (SEQ ID NO: 9), DP I-22.1 (SEQ IDNO: 29), and DP I-23.1 (SEQ ID NO: 33); DP I-1 (SEQ ID NO: 9), DP I-22.1(SEQ ID NO: 29), and DP I-25.2 (SEQ ID NO: 36); DP I-21.1 (SEQ ID NO:27), DP I-22.1 (SEQ ID NO: 29), DP I-23.1 (SEQ ID NO: 33), and DP I-25.2(SEQ ID NO: 36); DP I-21.2 (SEQ ID NO: 28), DP I-22.1 (SEQ ID NO: 29),and DP I-25.2 (SEQ ID NO: 36); DP I-21.2 (SEQ ID NO: 27), DP I-22.1 (SEQID NO: 29), DP I-25.2 (SEQ ID NO: 36), and DP I-26.1 (SEQ ID NO: 37); DFI-21.2 (SEQ ID NO: 91) and DF I-22.1 (SEQ ID NO: 92); DF I-21.1 (SEQ IDNO: 90), DF I-22.1 (SEQ ID NO: 92), and DF I-25.1 (SEQ ID NO: 97); DFI-21.2 (SEQ ID NO: 91), DF I-22.1 (SEQ ID NO: 92), and DF I-25.1 (SEQ IDNO: 97); DF I-1 (SEQ ID NO: 72) and DF I-22.1 (SEQ ID NO: 92); DF I-1(SEQ ID NO: 72), DF I-22.1 (SEQ ID NO: 92), and DF I-25.1 (SEQ ID NO:97); DF I-22.1 (SEQ ID NO: 29), and DF I-25.1 (SEQ ID NO: 35); DF I-21.1(SEQ ID NO: 90), DF I-22.1 (SEQ ID NO: 92), and DF I-23.1 (SEQ ID NO:95); DP I-21.1 (SEQ ID NO: 27), and DF I-22.1 (SEQ ID NO: 92); DP I-1(SEQ ID NO: 9), DP I-23.1 (SEQ ID NO: 33), DP I-25.1 (SEQ ID NO: 35),and DF I-1 (SEQ ID NO: 72); t) DP I-1 (SEQ ID NO: 9), DP I-25.1 (SEQ IDNO: 35), DP I-23.1 (SEQ ID NO: 33), and DF I-21.2 (SEQ ID NO: 91); DPI-1 (SEQ ID NO: 9), DP I-25.1 (SEQ ID NO: 35), DP I-23.1 (SEQ ID NO:33), and DF I-21.1 (SEQ ID NO: 90); DP II-22 (SEQ ID NO: 63), and DPII-25.2 (SEQ ID NO: 71); DP II-22 (SEQ ID NO: 63), DP II-25.2 (SEQ IDNO: 71), and DP I-21.1 (SEQ ID NO: 27) and DP I-22.1 (SEQ ID NO: 29); DPII-22 (SEQ ID NO: 63), DP II-25.2 (SEQ ID NO: 71), DP II-20.6 (SEQ IDNO: 56), DP I-22.1 (SEQ ID NO: 29), DP I-21.1 (SEQ ID NO: 27), and DPI-23.1 (SEQ ID NO: 33); DP II-22 (SEQ ID NO: 63), DP II-25.2 (SEQ ID NO:71), DP II-20.6 (SEQ ID NO: 56), DP I-21.1 (SEQ ID NO: 27), DP I-22.1(SEQ ID NO: 29), and DP I-25.2 (SEQ ID NO: 36); DP II-22 (SEQ ID NO:63), DP II-25.2 (SEQ ID NO: 71), DP I-21.1 (SEQ ID NO: 27), DP I-22.1(SEQ ID NO: 29), and DP I-25.2 (SEQ ID NO: 36); DP II-22 (SEQ ID NO:63), DP II-25.2 (SEQ ID NO: 71), DP I-21.1 (SEQ ID NO: 27), DP I-22.1(SEQ ID NO: 29), and DP I-23.1 (SEQ ID NO: 33); DP II-22 (SEQ ID NO:63), DP II-25.2 (SEQ ID NO: 71), DP I-1 (SEQ ID NO: 9), and DP I-22.1(SEQ ID NO: 29); DF II-4.5 (SEQ ID NO: 107) and DF II-2 (SEQ ID NO:104); DF II-4.5 (SEQ ID NO: 107) and DF II-19.1 (SEQ ID NO: 114); DFII-4.5 (SEQ ID NO: 107), DF II-2 (SEQ ID NO: 104), and DF II-19.1 (SEQID NO: 114); DF II-4.5 (SEQ ID NO: 107), DF II-2 (SEQ ID NO: 104), andDF II-9 (SEQ ID NO: 86); DF II-4.5 (SEQ ID NO: 107); and DF I-21.1 (SEQID NO: 90); DF II-4.5 (SEQ ID NO: 107), DP II-22 (SEQ ID NO: 63), and DPII-25.2 (SEQ ID NO: 71); and DF II-4.5 (SEQ ID NO: 107), DF II-2 (SEQ IDNO: 104), and DP II-22 (SEQ ID NO: 63).

Additional preferred peptides comprising a combination of two or moreRegions include the following combinations: DP I-21.2 (SEQ ID NO:28), DPI-23.1 (SEQ ID NO:33), DP I-26.1 (SEQ ID NO:37), DP II-20.6 (SEQ IDNO:56), DP II-22 (SEQ ID NO:63), DP II-25.2 (SEQ ID NO:71) and DPII-22.2 (SEQ ID NO:156); DP I-21.2 (SEQ ID NO:91); DP II-22.2 (SEQ IDNO:156); DP I-21.2 (SEQ ID NO:28), DP I-22.2 (SEQ ID NO:93), DP I-23.1(SEQ ID NO:95), DP I-25.2 (SEQ ID NO:36), DP I-26.1 (SEQ ID NO:37), DPI-27.1 (SEQ ID NO:38), DP II-20.6 (SEQ ID NO:56), DP II-22 (SEQ IDNO:63), DP II-24 (SEQ ID NO:68), and DP II-25.2; DP I-23.1, DP I-21.2,DP I-22, DF I-22.2, DP II-20.6, and DP II-25.2 (SEQ ID NO:71); DP I-23.1(SEQ ID NO:33), DP I-21.2 (SEQ ID NO:28), DP I-22.1 (SEQ ID NO:29), DFI-22.2 (SEQ ID NO:93); DP II-20.6 (SEQ ID NO:56), and DP II-25.2 (SEQ IDNO:71); DP I-23.1 (SEQ ID NO:33), DP I-21-2 (SEQ ID NO:28); DF I-22.2(SEQ ID NO:93) and DP II-20.6 (SEQ ID NO:56), and DP II-25.2 (SEQ IDNO:71); DP I-23.1 (SEQ ID NO:33), DF I-22.1 (SEQ ID NO:92), and DPII-20.6 (SEQ ID NO:56); DP I-26.1 (SEQ ID NO:37), DF I-22.2 (SEQ IDNO:93), and DP II-25.2 (SEQ ID NO:71); DP I-21.2 (SEQ ID NO:28), DFI-22.2 (SEQ ID NO:93), and DP II-22 (SEQ ID NO:63); and DP I-21.2 (SEQID NO:28) and DP II-22 (SEQ ID NO:63) all as shown in FIG. 29 and FIG.30.

Other preferred peptides comprise a combination of two or more regions(each region having an amino acid sequence as shown in FIG. 3 and FIG.36) derived from mite allergens Der p I, Der p II, and Der f I each ofsaid preferred peptides having the following specific sequentialarrangement of amino acid sequences as shown in FIGS. 25-27: DPI-26.1,DPII-25.2, DFI-22.2, DP II-22, DP I-23.1, DPII-20.6 and DPI-21.2 (SEQ IDNO:193) respectively; DPII-25.2, DFI-22.2, DPI-23.1, DPII-22, DPI-21.2and DPII-20.6 (SEQ ID NO:195) respectively; and DPII-25.2, DPI-21.2,DPI-23.1, DPI-26.1, DPII-22, DPII-20.6 and DFI-22.2 (SEQ ID NO:197)respectively. The nucleic acid and amino acid sequences of the abovepeptides are shown in FIGS. 25 (SEQ ID NOS:192 and 193), 26 (SEQ IDNOS:194 and 195) and 27 (SEQ ID NOS:196 and 197) respectively.

Other preferred peptides comprise various combinations of two or moreregions, each region comprising at least one T cell epitope of a proteinallergen of the genus Dermatophagoides, the regions may be derived fromthe same or different protein allergens of the genus Dermatophagoides,wherein at least one region comprises an amino acid sequence selectedfrom the following group: DP I-21.1 (SEQ ID NO: 27); DP I-21.2 (SEQ IDNO: 28); DP I-22.1 (SEQ ID NO: 29); DP I-23.1 (SEQ ID NO: 33); DP I-25.2(SEQ ID NO: 36); DP I-26.1 (SEQ ID NO: 37); DP I-28.1 (SEQ ID NO: 39);DP I-1 (SEQ ID NO: 9); DF I-1 (SEQ ID NO: 72); DF I-21.1 (SEQ ID NO:90); DF I-22.1 (SEQ ID NO: 92); DF I-23.1 (SEQ ID NO: 95); DF I-25.1(SEQ ID NO: 97); DP II-1 (SEQ ID NO: 41); DP II-1.2 (SEQ ID NO: 58); DPII-2.0 (SEQ ID NO: 56); DP II-20.3 (SEQ ID NO: 53); DP II-21 (SEQ ID NO:62); DP II-22 (SEQ ID NO: 63); DP II-25 (SEQ ID NO: 69); DP II-25.2 (SEQID NO: 71); DF II-2 (SEQ ID NO: 104); DF II-4.5 (SEQ ID NO: 107); DFII-15 (SEQ ID NO: 109); DF II-17 (SEQ ID NO: 111); DF II-19.1 (SEQ IDNO: 114); DF I-22.2 (SEQ ID NO:93); DP II-20.0 (SEQ ID NO: 50) and DPII-20.6 (SEQ ID NO:56), all as shown in FIGS. 3-4 and wherein at leastone region comprises an amino acid sequence selected from the followinggroup: DP I-21.7 (SEQ ID NO: 120); DP I-23.10 (SEQ ID NO: 121); DPI-23.11 (SEQ ID NO: 124); DP I-23.12 (SEQ ID NO: 125); DP I-23.5 (SEQ IDNO: 126); DP I-23.6 (SEQ ID NO: 127); DP I-23.7 (SEQ ID NO: 128); DPI-23.8 (SEQ ID NO: 129); DP I-23.9 (SEQ ID NO: 130); DP I-26.2 (SEQ IDNO: 134); DP II-20.7 (SEQ ID NO: 138); DP II-22.6 (SEQ ID NO: 139); DPII-22.3 (SEQ ID NO: 140); DP II-22.4 (SEQ ID NO: 141); DP II-22.5 (SEQID NO: 142); DP II-25.3 (SEQ ID NO: 148); DP II-25.4 (SEQ ID NO: 149);DP I-23.13 (SEQ ID NO: 122); DP I-23.14 (SEQ ID NO: 123); DP I-23.15(SEQ ID NO: 131); DP I-23.16 (SEQ ID NO: 132); DP I-23.17 (SEQ ID NO:133); DP I-26.3 (SEQ ID NO: 135); DP I-26.4 (SEQ ID NO: 136); DP I-26.5(SEQ ID NO: 137); DP II-22.7 (SEQ ID NO: 143); DP II-22.8 (SEQ ID NO:144); DP II-22.9 (SEQ ID NO: 145); DP II-22.10 (SEQ ID NO: 146); DPII-22.11 (SEQ ID NO: 147); DP I-23.32 (SEQ ID NO: 163), DPI-23.33 (SEQID NO: 164) DP I-23.31 (SEQ ID NO: 165), DP I-23.34 (SEQ ID NO: 166), DPI-23.35 (SEQ ID NO: 167), DP I-26.6 (SEQ ID NO: 168), DP II-20.9 (SEQ IDNO: 169), DP II-20.11 (SEQ ID NO: 169), DP II-20.10 (SEQ ID NO: 170) DPII-20.8 (SEQ ID NO: 171), D,P II-22.19 (SEQ ID NO: 172), DP II-22.20(SEQ ID NO: 173), DP II-22.21 (SEQ ID NO: 174), DP II-22.22 (SEQ ID NO:175), DP II-22.26 (SEQ ID NO: 176), DP II-22.23 (SEQ ID NO: 177), DPII-22.24 (SEQ ID NO: 178), DP II-22.25 (SEQ ID NO: 179), DP II-22.14(SEQ ID NO: 180), DF II-25.11 (SEQ ID NO: 182), DP II-25.9 (SEQ ID NO:183), DF II-25.10 (SEQ ID NO: 184), DF II-25.13 (SEQ ID NO: 186), DPII-25.14 (SEQ ID NO: 187), DP II-25.15 (SEQ ID NO: 188), DP II-25.16(SEQ ID NO: 189), DP II-25.17 (SEQ ID NO: 190), DP II-25.18 (SEQ ID NO:191) all as shown in FIGS. 29 and 30.

Additional preferred peptides comprise a combination of two or moreregions (each region having an amino acid sequence as shown in FIG. 3,FIG. 36, and FIG. 29) including: DP I-21.7 (SEQ ID NO:120), DF I-22.2(SEQ ID NO:93), DP I-23.13 (SEQ ID NO: 122), DP I-26.1 (SEQ ID NO:37),DP II-20.6 (SEQ ID NO:56), DP II-22.3 (SEQ ID NO: 140), and DP II-25.2(SEQ ID NO:71); DP I-21.2 (SEQ ID NO:28), DF I-22.2 (SEQ ID NO:93), DPI-23.10 (SEQ ID NO:121), DP I-26.2 (SEQ ID NO:134), DP II-20.6 (SEQ IDNO:56), DP II-22.4 (SEQ ID NO:141), and DP II-25.3 (SEQ ID NO: 148), DPI-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.11 (SEQ ID NO:124), DP I-26.3 (SEQ ID NO:135), DP II-20.7 (SEQ ID NO:138), DP II-22.5(SEQ ID NO: 142), and DP II-25.4 (SEQ ID NO:149); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.12 (SEQ ID NO: 125), DP I-26.4(SEQ ID NO: 136), DP II-20.6 (SEQ ID NO:56), DP II-22.6 (SEQ ID NO:139), and DP II-25.2 (SEQ ID NO:71); DP I-21.7 (SEQ ID NO:120), DFI-22.2 (SEQ ID NO:93), DP I-23.5 (SEQ ID NO: 126), DP I-26.5 (SEQ ID NO:137), DP II-20.7 (SEQ ID NO: 138), DP II-22.7 (SEQ ID NO: 143), and DPII-25.3 (SEQ ID NO: 148); DP I-21.7 (SEQ ID NO: 120), DF I-22.2 (SEQ IDNO:93), DP I-23.6 (SEQ ID NO: 127), DP I-26.2 (SEQ ID NO:134), DPII-20.6 (SEQ ID NO:56), DP II-22.8 (SEQ ID NO:144), and DP II-25.4 (SEQID NO: 149); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DPI-23.7 (SEQ ID NO: 128), DP I-26.3 (SEQ ID NO:135), DP II-20.7 (SEQ IDNO:138), DP II-22.9 (SEQ ID NO: 145), and DP II-25.3 (SEQ ID NO: 148);DP I-21.7 (SEQ ID NO: 120), DF I-22.2 (SEQ ID NO:93), DP I-23.8 (SEQ IDNO: 129), DP I-26.4 (SEQ ID NO: 136), DP II-20.6 (SEQ ID NO: 56), DPII-22.3 (SEQ ID NO: 140) and DP II-25.4 (SEQ ID NO: 149); DP I-21.7 (SEQID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.9 (SEQ ID NO:130), DPI-26.1 (SEQ ID NO:37), DP II-20.7 (SEQ ID NO: 138), DP II-22.4 (SEQ IDNO: 141), and DP II-25.3 (SEQ ID NO:148); DP I-21.7 (SEQ ID NO: 120), DFI-22.2 (SEQ ID NO:93), DP I-23.13 (SEQ ID NO:122), DP I-26.2 (SEQ ID NO:134), DP II-20.6 (SEQ ID NO:56), DP II-22.5 (SEQ ID NO: 142), and DPII-25.4 (SEQ ID NO:149); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ IDNO:93), DP I-23.14 (SEQ ID NO: 123), DP I-26.3 (SEQ ID NO: 135), DPII-20.7 (SEQ ID NO:138), DP II-22.6 (SEQ ID NO:139), and DP II-25.3 (SEQID NO:148); DP I-21.7 (SEQ ID NO: 120), DF I-22.2 (SEQ ID NO:93), DPI-23.15 (SEQ ID NO:131), DPI-26.4 (SEQ ID NO:136), DP II-20.6 (SEQ IDNO:56), DP II-22.6 (SEQ ID NO:139), and DP II-25.3 (SEQ ID NO: 148); DPI-21.7 (SEQ ID NO: 120), DF I-22.2 (SEQ ID NO:93), DP I-23.16 (SEQ IDNO:132), DP I-26.5 (SEQ ID NO:137), DP II-20.6 (SEQ ID NO:56), DPII-22.3 (SEQ ID NO:140), and DP II-25.4 (SEQ ID NO: 149); DP I-21.7 (SEQID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.17 (SEQ ID NO:133), DPI-26.2 (SEQ ID NO:134), DP II-20.6 (SEQ ID NO:56), DP II-22.4 (SEQ IDNO: 141), and DP II-25.3 (SEQ ID NO: 148); DP I-21.7 (SEQ ID NO:120), DFI-22.2 (SEQ ID NO:93), DP I-23.8 (SEQ ID NO:129), DP I-26.3 (SEQ IDNO:135), DP II-20.6 (SEQ ID NO:56), DP II-22.5 (SEQ ID NO: 142), and DPII-25.4 (SEQ ID NO:149); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ IDNO:93), DP I-23.9 (SEQ ID NO:130), DP I-26.4 (SEQ ID NO:136), DP II-20.6(SEQ ID NO:56), DP II-22.6 (SEQ ID NO:139) and DP II-25.3 (SEQ IDNO:148); and DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DPI-23.5 (SEQ ID NO:126), DP I-26.1 (SEQ ID NO:37), DP II-20.6 (SEQ IDNO:56), DP II-22.7 (SEQ ID NO:143), and DP II-25.4 (SEQ ID NO: 149).

Additional preferred peptides comprise at least two of the followingcombination of regions in any arrangement: X,Y,Z,A,B,C,D, wherein X isDP I-21.2 (SEQ ID NO: 27) or DP I-21.7 (SEQ ID NO: 120); Y is DF I-22.2(SEQ ID NO:93); Z is DP I-23.1 (SEQ ID NO:33), DP I-23.10 (SEQ IDNO:121), DP I-23.11 (SEQ ID NO:124), DP I-23.12 (SEQ ID NO:125), DPI-23.13 (SEQ ID NO: 122), DP I-23.14 (SEQ ID NO: 123), DP I-23.15 (SEQID NO:131), DP I-23.16 (SEQ ID NO:132), DP I-23.17 (SEQ ID NO:133), DPI-23.5 (SEQ ID NO:126), DP I-23.6 (SEQ ID NO:127), DP I-23.7 (SEQ IDNO:128), DP I-23.8 (SEQ ID NO: 129), DP I-23.9 (SEQ ID NO:130), DPI-23.32 (SEQ ID NO:163), DP I-23.33 (SEQ ID NO:164), DP I-23.31 (SEQ IDNO:165), DP I-23.34 (SEQ ID NO: 166), or DP I-23.35 (SEQ ID NO: 167); Ais DP I-26.1 (SEQ ID NO:37), DP I-26.2 (SEQ ID NO:134), DP I-26.3 (SEQID NO:135), DP I-26.4 (SEQ ID NO: 136), DP I-26.5 (SEQ ID NO: 137) or DPI 26.6 (SEQ ID NO:168); B is DP II-20.0 (SEQ ID NO:50), DP II-20.6 (SEQID NO:56), DP II-20.7 (SEQ ID NO:138), DP II-20.9 (SEQ ID NO:169), DPII-20.11 (SEQ ID NO:169), DP II-20.10 (SEQ ID NO: 170), or DP II-20.8(SEQ ID NO: 171); C is DP II-22 (SEQ ID NO:63), DP II-22.6 (SEQ ID NO:139), DP II-22.7 (SEQ ID NO: 143), DP II-22.8 (SEQ ID NO:144), DPII-22.9 (SEQ ID NO:145), DP II-22.10 (SEQ ID NO: 146), DP II-22.11 (SEQID NO: 147), DP II-22.3 (SEQ ID NO: 140), DP II-22.4 (SEQ ID NO:141), DPII-22.5 (SEQ ID NO:142), DP II-22.19 (SEQ ID NO:172), DP II-22.20 (SEQID NO:173), DP II-22.21 (SEQ ID NO:174), DP II-22.22 (SEQ ID NO:175), DPII-22.26 (SEQ ID NO:176), DP II-22.23 (SEQ ID NO: 177), DP II-22.24 (SEQID NO: 178), DP II-22.25 (SEQ ID NO: 179), or DP II-22.14 (SEQ IDNO:180); and D is DP II-25.2 (SEQ ID NO:71), DP II-25.3 (SEQ ID NO:148), DP II-25.4 (SEQ ID NO: 149), DF II-25.11 (SEQ ID NO: 182), DPII-25.9 (SEQ ID NO: 183), DF II-25.10 (SEQ ID NO: 184), or DF II-25.13(SEQ ID NO:186), DP II-25-14 (SEQ ID NO:187), DP II-25.15 (SEQ IDNO:188), DP II-25.16 (SEQ ID NO:189), DP II-25.17 (SEQ ID NO: 190), DPII-25.18 (SEQ ID NO: 191); with the proviso that X, Y, Z, A, B, C, D isnot the following combination of regions: DP I-21.2 (SEQ ID NO: 91), DFI-22.2 (SEQ ID NO: 93), DP I-23.1 (SEQ ID NO: 95), DP I-26.1 (SEQ ID NO:99), DP II-20.6 (SEQ ID NO: 56), DP II-22 (SEQ ID NO: 63), and DPII-25.2 (SEQ ID NO: 71). Preferably, a preferred peptide comprises atleast one region from each of X, Y, Z, A, B, C, and D.

Another preferred peptide comprises a specific sequential arrangement ofamino acid sequences, said specific sequential arrangement having theformula:

    ADYCZBX

wherein X is DP I-21.2 (SEQ ID NO: 27) or DP I-21.7 (SEQ ID NO:120); Yis DF I-22.2 (SEQ ID NO:93); Z is DP I-23.1 (SEQ ID NO:33), DP I-23.10(SEQ ID NO: 121), DP I-23.11 (SEQ ID NO:124), DP I-23.12 (SEQ IDNO:125), DP I-23.13 (SEQ ID NO:122), DP I-23.14 (SEQ ID NO:123), DPI-23.15 (SEQ ID NO:131), DP I-23.16 (SEQ ID NO:132), DP I-23.17 (SEQ IDNO:133), DP I-23.5 (SEQ ID NO:126), DP I-23.6 (SEQ ID NO:127), DP I-23.7(SEQ ID NO: 128), DP I-23.8 (SEQ ID NO: 129), DP I-23.9 (SEQ ID NO:130), DP I-23.32 (SEQ ID NO:163), DP I-23.33 (SEQ ID NO: 164), DPI-23.31 (SEQ ID NO:165), DP I-23.34 (SEQ ID NO:166), or DP I-23.35 (SEQID NO:167); A is DP I-26.1 (SEQ ID NO:37), DP I-26.2 (SEQ ID NO:134), DPI-26.3 (SEQ ID NO:135), DP I-26.4 (SEQ ID NO:136), DP I-26.5 (SEQ IDNO:137) or DP I 26.6 (SEQ ID NO: 168); B is DP II-20.0 (SEQ ID NO:50),DP II-20.6 (SEQ ID NO:56), DP II-20.7 (SEQ ID NO:138), DP II-20.9 (SEQID NO:169), DP II-20.11 (SEQ ID NO:169), DP II-20.10 (SEQ ID NO:170), orDP II-20.8 (SEQ ID NO:171); C is DP II-22 (SEQ ID NO:63), DP II-22.6(SEQ ID NO:139), DP II-22.7 (SEQ ID NO: 143), DP II-22.8 (SEQ IDNO:144), DP II-22.9 (SEQ ID NO: 145), DP II-22.10 (SEQ ID NO:146), DPII-22.11 (SEQ ID NO:147), DP II-22.3 (SEQ ID NO:140), DP II-22.4 (SEQ IDNO:141), DP II-22.5 (SEQ ID NO: 142), DP II-22.19 (SEQ ID NO:172), DPII-22.20 (SEQ ID NO:173), DP II-22.21 (SEQ ID NO: 174), DP II-22.22 (SEQID NO: 175), DP II-22.26 (SEQ ID NO: 176), DP II-22.23 (SEQ ID NO:177),DP II-22.24 (SEQ ID NO:178), DP II-22.25 (SEQ ID NO: 179), or DPII-22.14 (SEQ ID NO:180); and D is DP II-25.2 (SEQ ID NO:71), DP II-25.3(SEQ ID NO: 148), DP II-25.4 (SEQ ID NO: 149), DF II-25.11 (SEQ IDNO:182), DP II-25.9 (SEQ ID NO:183), DF II-25.10 (SEQ ID NO:184), or DFII-25.13 (SEQ ID NO:186), DP II-25-14 (SEQ ID NO:187), DP II-25.15 (SEQID NO:188), DP II-25.16 (SEQ ID NO:189), DP II-25.17 (SEQ ID NO: 190),DP II-25.18 (SEQ ID NO:191), with the proviso that ADYCZBX is not DPI-26.1 (SEQ ID NO: 37), DP II-25.2 (SEQ ID NO: 71), DF I-22.2 (SEQ IDNO: 93), DP II-22 (SEQ ID NO: 63), DP I-23.1 (SEQ ID NO: 95), DP II-20.6(SEQ ID NO: 56) and DP I-21.2 (SEQ ID NO:91) respectively.

Another preferred peptide comprises a specific sequenctial arrangementof amino acid sequences, said specific sequential arrangement having theformula:

    DYZCAXB

wherein X is DP I-21.2 (SEQ ID NO: 27) or DP I-21.7 (SEQ ID NO:120); Yis DF I-22.2 (SEQ ID NO:93); Z is DP I-23.1 (SEQ ID NO:33), DP I-23.10(SEQ ID NO:121), DP I-23.11 (SEQ ID NO:124), DP I-23.12 (SEQ ID NO:125),DP I-23.13 (SEQ ID NO:122), DP I-23.14 (SEQ ID NO:123), DP I-23.15 (SEQID NO:131), DP I-23.16 (SEQ ID NO:132), DP I-23.17 (SEQ ID NO:133),DPI-23.5 (SEQ ID NO:126), DP I-23.6 (SEQ ID NO:127), DP I-23.7 (SEQ IDNO:128), DP I-23.8 (SEQ ID NO:129), DP I-23.9 (SEQ ID NO:130), DPI-23.32 (SEQ ID NO:163), DP I-23.33 (SEQ ID NO:164), DP I-23.31 (SEQ IDNO:165), DP I-23.34 (SEQ ID NO:166), or DP I-23.35 (SEQ ID NO:167); A isDP I-26.1 (SEQ ID NO:37), DP I-26.2 (SEQ ID NO:134), DP I-26.3 (SEQ IDNO:135), DP I-26.4 (SEQ ID NO:136), DP I-26.5 (SEQ ID NO:137) or DPI-26.6 (SEQ ID NO: 168); B is DP II-20.0 (SEQ ID NO:50), DP II-20.6 (SEQID NO:56), DP II-20.7 (SEQ ID NO:138), DP II-20.9 (SEQ ID NO:169), DPII-20.11 (SEQ ID NO: 169), DP II-20.10 (SEQ ID NO:170), or DP II-20.8(SEQ ID NO:171); C is DP II-22 (SEQ ID NO:63), DP II-22.6 (SEQ ID NO:139), DP II-22.7 (SEQ ID NO: 143), DP II-22.8 (SEQ ID NO: 144), DPII-22.9 (SEQ ID NO: 145), DP II-22.10 (SEQ ID NO: 146), DP II-22.11 (SEQID NO: 147), DP II-22.3 (SEQ ID NO: 140), DP II-22.4 (SEQ ID NO: 141),DP II-22.5 (SEQ ID NO: 142), DP II-22.19 (SEQ ID NO: 172), DP II-22.20(SEQ ID NO: 173), DP II-22.21 (SEQ ID NO: 174), DP II-22.22 (SEQ ID NO:175), DP II-22.26 (SEQ ID NO: 176), DP II-22.23 (SEQ ID NO: 177), DPII-22.24 (SEQ ID NO:178), DP II-22.25 (SEQ ID NO:179), or DP II-22.14(SEQ ID NO:180); and D is DP II-25.2 (SEQ ID NO:71), DP II-25.3 (SEQ IDNO: 148), DP II-25.4 (SEQ ID NO: 149), DF II-25.11 (SEQ ID NO:182), DPII-25.9 (SEQ ID NO: 183), DF II-25.10 (SEQ ID NO: 184), or DF II-25.13(SEQ ID NO: 186), DP II-25-14 (SEQ ID NO: 187), DP II-25.15 (SEQ IDNO:188), DP II-25.16 (SEQ ID NO:189), DP II-25.17 (SEQ ID NO: 190), DPII-25.18 (SEQ ID NO:191); with the proviso that DYZCAXB is not DPII-25.2 (SEQ ID NO: 71) , DF I-22.2 (SEQ ID NO: 93), DP I-23.1 (SEQ IDNO: 95), DP II-22 (SEQ ID NO: 63), DP I-26.1 (SEQ ID NO: 37), DP I-21.2(SEQ ID NO: 91) and DP II-20.6 (SEQ ID NO: 56) respectively.

Yet another preferred peptide has a specific arrangement of amino acidsequences, said specific sequential arrangement having the formula:

    DXZACBY

wherein X is DP I-21.2 (SEQ ID NO: 27) or DP I-21.7 (SEQ ID NO:120); Yis DF I-22.2 (SEQ ID NO:93); Z is DP I-23.1 (SEQ ID NO:33), DP I-23.10(SEQ ID NO:121), DP I-23.11 (SEQ ID NO:124), DP I-23.12 (SEQ ID NO:125),DP I-23.13 (SEQ ID NO:122), DP I-23.14 (SEQ ID NO:123), DP I-23.15 (SEQID NO: 131), DP I-23.16 (SEQ ID NO:132), DP I-23.17 (SEQ ID NO: 133), DPI-23.5 (SEQ ID NO:126), DP I-23.6 (SEQ ID NO:127), DP I-23.7 (SEQ ID NO:128), DP I-23.8 (SEQ ID NO: 129), DP I-23.9 (SEQ ID NO:130), DP I-23.32(SEQ ID NO: 163), DP I-23.33 (SEQ ID NO: 164), DP I-23.31 (SEQ ID NO:165), DP I-23.34 (SEQ ID NO:166), or DP I-23.35 (SEQ ID NO:167); A is DPI-26.1 (SEQ ID NO: 37), DP I-26.2 (SEQ ID NO: 134), DP I-26.3 (SEQ IDNO: 135), DP I-26.4 (SEQ ID NO: 136), DP I-26.5 (SEQ ID NO:137) or DPI-26.6 (SEQ ID NO:168); B is DP II-20.0 (SEQ ID NO:50), DP II-20.6 (SEQID NO:56), DP II-20.7 (SEQ ID NO: 138), DP II-20.9 (SEQ ID NO: 169), DPII-20.11 (SEQ ID NO: 169), DP II-20.10 (SEQ ID NO:170), or DP II-20.8(SEQ ID NO:171); C is DP II-22 (SEQ ID NO:63), DP II-22.6 (SEQ IDNO:139), DP II-22.7 (SEQ ID NO: 143), DP II-22.8 (SEQ ID NO: 144), DPII-22.9 (SEQ ID NO: 145), DP II-22.10 (SEQ ID NO: 146), DP II-22.11 (SEQID NO: 147), DP II-22.3 (SEQ ID NO: 140), DP II-22.4 (SEQ ID NO: 141),DP II-22.5 (SEQ ID NO: 142), DP II-22.19 (SEQ ID NO: 172), DP II-22.20(SEQ ID NO: 173), DP II-22.21 (SEQ ID NO: 174), DP II-22.22 (SEQ ID NO:175), DP II-22.26 (SEQ ID NO: 176), DP II-22.23 (SEQ ID NO: 177), DPII-22.24 (SEQ ID NO: 178), DP II-22.25 (SEQ ID NO: 179), or DP II-22.14(SEQ ID NO:180); and D is DP II-25.2 (SEQ ID NO:71), DP II-25.3 (SEQ IDNO: 148), DP II-25.4 (SEQ ID NO:149), DF II-25.11 (SEQ ID NO: 182), DPII-25.9 (SEQ ID NO: 183), DF II-25.10 (SEQ ID NO:184), or DF II-25.13(SEQ ID NO: 186), DP II-25-14 (SEQ ID NO:187), DP II-25.15 (SEQ IDNO:188), DP II-25.16 (SEQ ID NO:189), DP II-25.17 (SEQ ID NO: 190), DPII-25.18 (SEQ ID NO: 191); with the proviso that DXZACBY is not DPII-25.2 (SEQ ID NO: 71), DP I-21.2 (SEQ ID NO: 91), DP I-23.1 (SEQ IDNO: 95), DP I-26.1 (SEQ ID NO: 37), DP II-22 (SEQ ID NO: 63), DP II-20.6(SEQ ID NO: 56), and DF I-22.2 (SEQ ID NO: 93) respectively.

In yet another aspect of the present invention, a composition isprovided comprising at least two peptides (e.g., a physical mixture ofat least two peptides), each comprising at least one T cell epitope of aprotein allergen of the genus Dermatophagoides. The peptides or modifiedpeptides may be derived from the same or from different mite allergens.Such compositions can be administered in the form of a therapeuticcomposition with a pharmaceutically acceptable carrier of diluent. Atherapeutically effective amount of one or more of such compositions canbe administered simultaneously or sequentially to an individualsensitive to house dust mite. In addition, peptides derived from thesame or different mite allergens can be administered simultaneously orsequentially. Such combinations of peptides may comprise therapeuticcompositions comprising only one peptide or more peptides if desired.Such compositions may be administered simultaneously or sequentially inpreferred combinations.

Preferred compositions and preferred combinations of peptides which canbe administered simultaneously or sequentially (comprising peptideshaving amino acid sequences shown in FIG. 3 and FIG. 36) include thefollowing combinations: DP I-22.1 (SEQ ID NO: 29) and DP I-25.1 (SEQ IDNO: 35); DP I-21.1 (SEQ ID NO: 27) and DP I-25.2 (SEQ ID NO: 36); DPI-22.1 (SEQ ID NO: 29) and DP I-1 (SEQ ID NO: 9); DP I-21. I (SEQ ID NO:27), DP I-22.1 (SEQ ID NO: 29), and DP I-25.2 (SEQ ID NO: 36); DP I-21.2(SEQ ID NO: 28), DP I-22.1 (SEQ ID NO: 29), and DP I-23.1 (SEQ ID NO:39); DP I-1(SEQ ID NO: 9), DP I-22.1 (SEQ ID NO: 29), and DP I-23.1 (SEQID NO: 33); DP I-1 (SEQ ID NO: 9), DP I-22.1 (SEQ ID NO: 29), and DPI-25.2 (SEQ ID NO: 36); DP I-21.1 (SEQ ID NO: 27), DP I-22.1 (SEQ ID NO:29), DP I-23.1 (SEQ ID NO: 33), and DP I-25.2 (SEQ ID NO: 36); DP I-21.2(SEQ ID NO: 28), DP I-22.1 (SEQ ID NO: 29), and DP I-25.2 (SEQ ID NO:36); DP I-21.2 (SEQ ID NO: 27), DP I-22.1 (SEQ ID NO: 29), DP I-25.2(SEQ ID NO: 36), and DP I-26.1 (SEQ ID NO: 37); DF I-21.2 (SEQ ID NO:91) and DF I-22.1 (SEQ ID NO: 92); DF I-21.1 (SEQ ID NO: 90), DF I-22.1(SEQ ID NO: 92), and DF I-25.1 (SEQ ID NO: 97); DF I-21.2 (SEQ ID NO:91), DF I-22.1 (SEQ ID NO: 92), and DF I-25.1 (SEQ ID NO: 97); DF I-1(SEQ ID NO: 72) and DF I-22.1 (SEQ ID NO: 92); DF I-1 (SEQ ID NO: 72),DF I-22.1 (SEQ ID NO: 92), and DF I-25.1 (SEQ ID NO: 97); DF I-22.1 (SEQID NO: 29), and DF I-25.1 (SEQ ID NO: 35); DF I-21.1 (SEQ ID NO: 90), DFI-22.1 (SEQ ID NO: 92), and DF I-23.1 (SEQ ID NO: 95); DP I-21.1 (SEQ IDNO: 27), and DF I-22.1 (SEQ ID NO: 92); DP I-1 (SEQ ID NO: 9), DP I-23.1(SEQ ID NO: 33), DP I-25.1 (SEQ ID NO: 35), and DF I-1 (SEQ ID NO: 72);DP I-1 (SEQ ID NO: 9), DP I-25.1 (SEQ ID NO: 35), DP I-23.1 (SEQ ID NO:33), and DF I-21.2 (SEQ ID NO: 91); DP I-1 (SEQ ID NO: 9), DP I-25.1(SEQ ID NO: 35), DP I-23.1 (SEQ ID NO: 33), and DF I-21.1 (SEQ ID NO:90); DP II-22 (SEQ ID NO: 63), and DP II-25.2 (SEQ ID NO: 71); DP II-22(SEQ ID NO: 63), DP II-25.2 (SEQ ID NO: 71), and DP I-21.1 (SEQ ID NO:27) and DP I-22.1 (SEQ ID NO: 29); DP II-22 (SEQ ID NO: 63), DP II-25.2(SEQ ID NO: 71), DP II-20.6 (SEQ ID NO: 56), DP I-22.1 (SEQ ID NO: 29),DP I-21.1 (SEQ ID NO: 27), and DP I-23.1 (SEQ ID NO: 33); DP II-22 (SEQID NO: 63), DP II-25.2 (SEQ ID NO: 71), DP II-20.6 (SEQ ID NO: 56), DPI-21.1 (SEQ ID NO: 27), DP I-22.1 (SEQ ID NO: 29), and DP I-25.2 (SEQ IDNO: 36); DP II-22 (SEQ ID NO: 63), DP II-25.2 (SEQ ID NO: 71), DP I-21.1(SEQ ID NO: 27), DP I-22.1 (SEQ ID NO: 29), and DP I-25.2 (SEQ ID NO:36); DP II-22 (SEQ ID NO: 63), DP II-25.2 (SEQ ID NO: 71), DP I-21.1(SEQ ID NO: 27), DP I-22.1 (SEQ ID NO: 29), and DP I-23.1 (SEQ ID NO:33); DP II-22 (SEQ ID NO: 63), DP II-25.2 (SEQ ID NO: 71), DP I-1 (SEQID NO: 9), and DP I-22.1 (SEQ ID NO: 29); DF II-4.5 (SEQ ID NO: 107) andDF II-2 (SEQ ID NO: 104); DF II-4.5 (SEQ ID NO: 107) and DF II-19.1 (SEQID NO: 114); DF II-4.5 (SEQ ID NO: 107), DF II-2 (SEQ ID NO: 104), andDF II-19.1 (SEQ ID NO: 114); DF II-4.5 (SEQ ID NO: 107), DF II-2 (SEQ IDNO: 104), and DF II-9 (SEQ ID NO: 86); DF II-4.5 (SEQ ID NO: 107); andDF I-21.1 (SEQ ID NO: 90); DF II-4.5 (SEQ ID NO: 107), DP II-22 (SEQ IDNO: 63), and DP II-25.2 (SEQ ID NO: 71); and DF II-4.5 (SEQ ID NO: 107),DF II-2 (SEQ ID NO: 104), and DP II-22 (SEQ ID NO: 63); and DP I-26.1(SEQ ID NO: 65), DP II-25.2 (SEQ ID NO: 71), DF I-22 (SEQ ID NO: 63), DPII-20.6 (SEQ ID NO: 56) and DP I-21.2 (SEQ ID NO: 91).

Other preferred therapeutic compositions comprising a pharmaceuticallyacceptable carrier or diluent and at least two peptides each comprisingat least one T cell epitope, comprise at least one peptide selected fromthe group consisting of: DP I-1 (SEQ ID NO: 9); DP I-2 (SEQ ID NO: 10);DP I-3 (SEQ ID NO: 11); DP I-4 (SEQ ID NO: 12); DP I-11.1 (SEQ ID NO:13); DP I-12.1 (SEQ ID NO: 14); DP I-5 (SEQ ID NO: 15); DP I-13 (SEQ IDNO: 17); DP I-14 (SEQ ID NO: 18); DP I-15 (SEQ ID NO: 19); DP I-6.1 (SEQID NO: 20); DP I-7.1 (SEQ ID NO: 21); DP I-8 (SEQ ID NO: 22); DP I-9(SEQ ID NO: 23); DP I-16 (SEQ ID NO: 24); DP I-10 (SEQ ID NO: 25); DPI-17 (SEQ ID NO: 26); DP I-21.1 (SEQ ID NO: 27); DP I-21.2 (SEQ ID NO:28); DP I-22.1 (SEQ ID NO: 29); DP I-22.2 (SEQ ID NO: 30); DP I-22.3(SEQ ID NO: 31); DP I-22.4 (SEQ ID NO: 32); DP I-23.1 (SEQ ID NO: 33);DP I-23.2 (SEQ ID NO: 34); DP I-25.1 (SEQ ID NO: 35); DP I-25.2 (SEQ IDNO: 36); DP I-26.1 (SEQ ID NO: 37); DP I-27.1 (SEQ ID NO: 38); DP I-28.1(SEQ ID NO: 39); DP I-28.2 (SEQ ID NO: 40), DF I-1 (SEQ ID NO: 72); DFI-2.1 (SEQ ID NO: 73); DF I-3 (SEQ ID NO: 74); DF I-4 (SEQ ID NO: 75);DF I-11 (SEQ ID NO: 76); DF I-12 (SEQ ID NO: 77); DF I-5 (SEQ ID NO:78); DF I-13 (SEQ ID NO: 79); DF I-14 (SEQ ID NO: 80); DF I-15 (SEQ IDNO: 81); DF I-6 (SEQ ID NO: 82); DF I-7 (SEQ ID NO: 83); DF I-8.1 (SEQID NO: 84); DF I-8 (SEQ ID NO: 85); DF I-9 (SEQ ID NO: 86); DF I-16 (SEQID NO: 87); DF I-10 (SEQ ID NO: 88); DF I-17 (SEQ ID NO: 89); DF I-21.1(SEQ ID NO: 90); DF I-21.2 (SEQ ID NO: 91); DF I-22.1 (SEQ ID NO: 92);DF I-22.2 (SEQ ID NO: 93); DF I-22.4 (SEQ ID NO: 94); DF I-23.1 (SEQ IDNO: 95); DF I-23.2 (SEQ ID NO: 96); DF I-25.1 (SEQ ID NO: 97); DF I-25.2(SEQ ID NO: 98); DF I-26.1 (SEQ ID NO: 99); DF I-27.1 (SEQ ID NO: 100);DF I-28.1 (SEQ ID NO: 101); DF I-28.2 (SEQ ID NO: 102); DP II-20 (SEQ IDNO: 50); DP II-20.1 (SEQ ID NO: 51); DP II-20.2 (SEQ ID NO: 52); DPII-20.3 (SEQ ID NO: 53); DP II-20.4 (SEQ ID NO: 54); DP II-20.5 (SEQ IDNO: 55); DP II 20.6 (SEQ ID NO: 56); DP II-1 (SEQ ID NO: 41); DP II-2(SEQ ID NO: 42); DP II-3.1 (SEQ ID NO: 43); DP II-4 (SEQ ID NO: 44); DPII-5 (SEQ ID NO: 45); DP II-6 (SEQ ID NO: 46); DP II-7 (SEQ ID NO: 47);DP II-8 (SEQ ID NO: 48); DP II-9 (SEQ ID NO: 49); DP II-1.1 (SEQ ID NO:57); DP II-1.2 (SEQ ID NO: 58); DP II-2.1 (SEQ ID NO: 59); DP II-2.2(SEQ ID NO: 60); DP II-2.3 (SEQ ID NO: 61); DP II-21 (SEQ ID NO: 62); DPII-22 (SEQ ID NO: 63); DP II-26 (SEQ ID NO: 64); DP II-26.1 (SEQ ID NO:65); DP II-23 (SEQ ID NO: 66); DP II-23.1 (SEQ ID NO: 67); DP II-24 (SEQID NO: 68); DP II-25 (SEQ ID NO: 69); DP II-25.1 (SEQ ID NO: 70); DPII-25.2 (SEQ ID NO: 71); DF II-1 (SEQ ID NO: 103); DF II-2 (SEQ ID NO:104); DF II-13.1 (SEQ ID NO: 105); DF II-3.1 (SEQ ID NO: 106); DF II-4.5(SEQ ID NO: 107); DF II-4.3 (SEQ ID NO: 108); DF II-15 (SEQ ID NO: 109);DF II-16 (SEQ ID NO: 110); DF II-17 (SEQ ID NO: 111); DF II-18 (SEQ IDNO: 112); DF II-19 (SEQ ID NO: 113); DF II-19.1 (SEQ ID NO: 114); DFII-21 (SEQ ID NO: 115); and DF II-22 (SEQ ID NO: 116), DP I-23.1.1, DPI-23.1.2, DP I-23.1.3, DP I-23.1.4, DP II-22.1, II-22.2 (all as shown inFIGS. 3, 4, and 25), and at least one modified peptide selected from thefollowing group:DP I-21.7 (SEQ ID NO: 120); DP I-23.10 (SEQ ID NO: 121);DP I-23.11 (SEQ ID NO: 124); DP I-23.12 (SEQ ID NO: 125); DP I-23.5 (SEQID NO: 126); DP I-23.6 (SEQ ID NO: 127); DP I-23.7 (SEQ ID NO: 128); DPI-23.8 (SEQ ID NO: 129); DP I-23.9 (SEQ ID NO: 130); DP I-26.2 (SEQ IDNO: 134); DP II-20.7 (SEQ ID NO: 138); DP II-22.6 (SEQ ID NO: 139); DPII-22.3 (SEQ ID NO: 140); DP II-22.4 (SEQ ID NO: 141); DP II-22.5 (SEQID NO: 142); DP II-25.3 (SEQ ID NO: 148); DP II-25.4 (SEQ ID NO: 149);DP I-23.13 (SEQ ID NO: 122); DP I-23.14 (SEQ ID NO: 123); DP I-23.15(SEQ ID NO: 131); DP I-23.16 (SEQ ID NO: 132); DP I-23.17 (SEQ ID NO:133); DP I-26.3 (SEQ ID NO: 135); DP I-26.4 (SEQ ID NO: 136); DP I-26.5(SEQ ID NO: 137); DP II-22.7 (SEQ ID NO: 143); DP II-22.8 (SEQ ID NO:144); DP II-22.9 (SEQ ID NO: 145); DP II-22.10 (SEQ ID NO: 146); DPII-22.11 (SEQ ID NO: 147); DP I-23.32 (SEQ ID NO: 163), DPI-23.33 (SEQID NO: 164) DP I-23.31 (SEQ ID NO: 165), DP I-23.34 (SEQ ID NO: 166), DPI-23.35 (SEQ ID NO: 167), DP I-26.6 (SEQ ID NO: 168), DP II-20.9 (SEQ IDNO: 169), DP II-20.11 (SEQ ID NO: 169), DP II-20.10 (SEQ ID NO: 170) DPII-20.8 (SEQ ID NO: 171), D,P II-22.19 (SEQ ID NO: 172), DP II-22.20(SEQ ID NO: 173), DP II-22.21 (SEQ ID NO: 174), DP II-22.22 (SEQ ID NO:175), DP II-22.26 (SEQ ID NO: 176), DP II-22.23 (SEQ ID NO: 177), DPII-22.24 (SEQ ID NO: 178), DP II-22.25 (SEQ ID NO: 179), DP II-22.14(SEQ ID NO: 180), DF II-25.11 (SEQ ID NO: 182), DP II-25.9 (SEQ ID NO:183), DF II-25.10 (SEQ ID NO: 184), DF II-25.13 (SEQ ID NO: 186), DPII-25.14 (SEQ ID NO: 187), DP II-25.15 (SEQ ID NO: 188), DP II-25.16(SEQ ID NO: 189), DP II-25.17 (SEQ ID NO: 190), DP II-25.18 (SEQ ID NO:191) all as shown in FIGS. 29 and 30.

Even more preferred therapeutic compositions comprising apharmaceutically acceptable carrier or diluent and at least two peptideseach comprising at least one T cell epitope, comprise at least onepeptide selected from the group consisting of: DP I-21.7 (SEQ ID NO:120); DP I-23.10 (SEQ ID NO: 121); DP I-23.11 (SEQ ID NO: 124); DPI-23.12 (SEQ ID NO: 125); DP I-23.5 (SEQ ID NO: 126); DP I-23.6 (SEQ IDNO: 127); DP I-23.7 (SEQ ID NO: 128); DP I-23.8 (SEQ ID NO: 129); DPI-23.9 (SEQ ID NO: 130); DP I-26.2 (SEQ ID NO: 134); DP II-20.7 (SEQ IDNO: 138); DP II-22.6 (SEQ ID NO: 139); DP II-22.3 (SEQ ID NO: 140); DPII-22.4 (SEQ ID NO: 141); DP II-22.5 (SEQ ID NO: 142); DP II-25.3 (SEQID NO: 148); DP II-25.4 (SEQ ID NO: 149); DP I-23.13 (SEQ ID NO: 122);DP I-23.14 (SEQ ID NO: 123); DP I-23.15 (SEQ ID NO: 131); DP I-23.16(SEQ ID NO: 132); DP I-23.17 (SEQ ID NO: 133); DP I-26.3 (SEQ ID NO:135); DP I-26.4 (SEQ ID NO: 136); DP I-26.5 (SEQ ID NO: 137); DP II-22.7(SEQ ID NO: 143); DP II-22.8 (SEQ ID NO: 144); DP II-22.9 (SEQ ID NO:145); DP II-22.10 (SEQ ID NO: 146); DP II-22.11 (SEQ ID NO: 147); DPI-23.32 (SEQ ID NO: 163), DPI-23.33 (SEQ ID NO: 164) DP I-23.31 (SEQ IDNO: 165), DP I-23.34 (SEQ ID NO: 166), DP I-23.35 (SEQ ID NO: 167), DPI-26.6 (SEQ ID NO: 168), DP II-20.9 (SEQ ID NO: 169), DP II-20.11 (SEQID NO: 169), DP II-20.10 (SEQ ID NO: 170) DP II-20.8 (SEQ ID NO: 171),D,P II-22.19 (SEQ ID NO: 172), DP II-22.20 (SEQ ID NO: 173), DP II-22.21(SEQ ID NO: 174), DP II-22.22 (SEQ ID NO: 175), DP II-22.26 (SEQ ID NO:176), DP II-22.23 (SEQ ID NO: 177), DP II-22.24 (SEQ ID NO: 178), DPII-22.25 (SEQ ID NO: 179), DP II-22.14 (SEQ ID NO: 180), DF II-25.11(SEQ ID NO: 182), DP II-25.9 (SEQ ID NO: 183), DF II-25.10 (SEQ ID NO:184), DF II-25.13 (SEQ ID NO: 186), DP II-25.14 (SEQ ID NO: 187), DPII-25.15 (SEQ ID NO: 188), DP II-25.16 (SEQ ID NO: 189), DP II-25.17(SEQ ID NO: 190), DP II-25.18 (SEQ ID NO: 191) all as shown in FIGS. 29and 30, wherein said composition comprises a sufficient percentage ofthe T cell epitopes of at least one protein allergen such that uponadministration of the composition to an individual sensitive to a housedust mite allergen, T cells of the individual are tolerized to said atleast one protein allergen.

Other preferred compositions of the invention comprise the followingcombination of peptides and/or modified peptides and a pharmaceuticallyacceptable carrier or diluent: DP I-21.7 (SEQ ID NO: 120), DF I-22.2(SEQ ID NO:93), DP I-23.13 (SEQ ID NO:122), DP I-26.1 (SEQ ID NO:37), DPII-20.6 (SEQ ID NO:56), DP II-22.3 (SEQ ID NO:140), and DP II-25.2 (SEQID NO:71); DP I-21.2 (SEQ ID NO:28), DF I-22.2 (SEQ ID NO:93), DPI-23.10 (SEQ ID NO:121), DP I-26.2 (SEQ ID NO:134), DP I-20.6 (SEQ IDNO:56), DP II-22.4 (SEQ ID NO:141), and DP II-25.3 (SEQ ID NO: 148), DPI-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.11 (SEQ ID NO:124), DP I-26.3 (SEQ ID NO: 135), DP II-20.7 (SEQ ID NO: 138), DPII-22.5 (SEQ ID NO: 142), and DP II-25.4 (SEQ ID NO:149); DP I-21.7 (SEQID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.12 (SEQ ID NO:125), DPI-26.4 (SEQ ID NO:136), DP II-20.6 (SEQ ID NO:56), DP II-22.6 (SEQ IDNO:139), and DP II-25.2 (SEQ ID NO:71); DP I-21.7 (SEQ ID NO:120), DFI-22.2 (SEQ ID NO:93), DP I-23.5 (SEQ ID NO:126), DP I-26.5 (SEQ IDNO:137), DP II-20.7 (SEQ ID NO:138), DP II-22.7 (SEQ ID NO:143), and DPII-25.3 (SEQ ID NO:148); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ IDNO:93), DP I-23.6 (SEQ ID NO:127), DP I-26.2 (SEQ ID NO:134), DP II-20.6(SEQ ID NO:56), DP II-22.8 (SEQ ID NO: 144), and DP II-25.4 (SEQ IDNO:149); DP I-21.7 (SEQ ID NO: 120), DF I-22.2 (SEQ ID NO:93), DP I-23.7(SEQ ID NO:128), DP I-26.3 (SEQ ID NO:135), DP II-20.7 (SEQ ID NO:138),DP II-22.9 (SEQ ID NO:145), and DP II-25.3 (SEQ ID NO:148); DP I-21.7(SEQ ID NO: 120), DF I-22.2 (SEQ ID NO:93), DP I-23.8 (SEQ ID NO:129),DP I-26.4 (SEQ ID NO:136), DP II-20.6 (SEQ ID NO:56), DP II-22.3 (SEQ IDNO: 140) and DP II-25.4 (SEQ ID NO:149); DP I-21.7 (SEQ ID NO:120), DFI-22.2 (SEQ ID NO:93), DP I-23.9 (SEQ ID NO:130), DP I-26.1 (SEQ IDNO:37), DP II-20.7 (SEQ ID NO:138), DP II-22.4 (SEQ ID NO:141), and DPII-25.3 (SEQ ID NO:148); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ IDNO:93), DP I-23.13 (SEQ ID NO:122), DP I-26.2 (SEQ ID NO:134), DPII-20.6 (SEQ ID NO:56), DP II-22.5 (SEQ ID NO:142), and DP II-25.4 (SEQID NO:149); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DPI-23.14 (SEQ ID NO:123), DP I-26.3 (SEQ ID NO:135), DP II-20.7 (SEQ IDNO:138), DP II-22.6 (SEQ ID NO:139), and DP II-25.3 (SEQ ID NO:148); DPI-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.15 (SEQ IDNO:131), DP I-26.4 (SEQ ID NO:136), DP II-20.6 (SEQ ID NO:56), DPII-22.6 (SEQ ID NO:139), and DP II-25.3 (SEQ ID NO:148); DP I-21.7 (SEQID NO: 120), DF I-22.2 (SEQ ID NO:93), DP I-23.16 (SEQ ID NO:132), DPI-26.5 (SEQ ID NO: 137), DP II-20.6 (SEQ ID NO:56), DP II-22.3 (SEQ IDNO:140), and DP II-25.4 (SEQ ID NO: 149); DP I-21.7 (SEQ ID NO:120), DFI-22.2 (SEQ ID NO:93), DP I-23.17 (SEQ ID NO:133), DP I-26.2 (SEQ IDNO:134), DP II-20.6 (SEQ ID NO:56), DP II-22.4 (SEQ ID NO: 141), and DPII-25.3 (SEQ ID NO:148); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ IDNO:93), DP I-23.8 (SEQ ID NO:129), DP I-26.3 (SEQ ID NO:135), DP II-20.6(SEQ ID NO:56), DP II-22.5 (SEQ ID NO:142), and DP II-25.4 (SEQ IDNO:149); DP I-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.9(SEQ ID NO: 130), DP I-26.4 (SEQ ID NO: 136), DP II-20.6 (SEQ ID NO:56),DP II-22.6 (SEQ ID NO:139) and DP II-25.3 (SEQ ID NO: 148); and DPI-21.7 (SEQ ID NO:120), DF I-22.2 (SEQ ID NO:93), DP I-23.5 (SEQ ID NO:126), DP I-26.1 (SEQ ID NO:37), DP II-20.6 (SEQ ID NO:56), DP II-22.7(SEQ ID NO:143), and DP II-25.4 (SEQ ID NO:149).

In yet another aspect of the invention a composition is providedcomprising a pharmaceutically acceptable carrier or diluent and at leasttwo, preferably at least four, more preferably at least six, even morepreferably at least seven of the following combination of peptides: X,Y, Z, A, B, C, D, wherein X is DP I-21.2 (SEQ ID NO: 27) or DP I-21.7(SEQ ID NO:120); Y is DF I-22.2 (SEQ ID NO:93); Z is DP I-23.1 (SEQ IDNO:33), DP I-23.10 (SEQ ID NO:121), DP I-23.11 (SEQ ID NO:124), DPI-23.12 (SEQ ID NO:125), DP I-23.13 (SEQ ID NO:122), DP I-23.14 (SEQ IDNO:123), DP I-23.15 (SEQ ID NO:131), DP I-23.16 (SEQ ID NO:132), DPI-23.17 (SEQ ID NO:133), DP I-23.5 (SEQ ID NO:126), DP I-23.6 (SEQ IDNO:127), DP I-23.7 (SEQ ID NO:128), DP I-23.8 (SEQ ID NO:129), DP I-23.9(SEQ ID NO:130), DP I-23.32 (SEQ ID NO:163), DP I-23.33 (SEQ ID NO:164), DP I-23.31 (SEQ ID NO:165), DP I-23.34 (SEQ ID NO:166), or DPI-23.35 (SEQ ID NO:167); A is DP I-26.1 (SEQ ID NO:37), DP I-26.2 (SEQID NO:134), DP I-26.3 (SEQ ID NO:135), DP I-26.4 (SEQ ID NO:136), DPI-26.5 (SEQ ID NO:137) or DP I 26.6 (SEQ ID NO: 168); B is DP II-20.0(SEQ ID NO:50), DP II-20.6 (SEQ ID NO:56), DP II-20.7 (SEQ ID NO: 138),DP II-20.9 (SEQ ID NO: 169), DP II-20.11 (SEQ ID NO: 169), DP II-20.10(SEQ ID NO: 170), or DP II-20.8 (SEQ ID NO: 171); C is DP II-22 (SEQ IDNO:63), DP II-22.6 (SEQ ID NO:139), DP II-22.7 (SEQ ID NO:143), DPII-22.8 (SEQ ID NO: 144), DP II-22.9 (SEQ ID NO:145), DP II-22.10 (SEQID NO:146), DP II-22.11 (SEQ ID NO:147), DP II-22.3 (SEQ ID NO: 140), DPII-22.4 (SEQ ID NO:141), DP II-22.5 (SEQ ID NO:142), DP II-22.19 (SEQ IDNO:172), DP II-22.20 (SEQ ID NO:173), DP II-22.21 (SEQ ID NO: 174), DPII-22.22 (SEQ ID NO:175), DP II-22.26 (SEQ ID NO:176), DP II-22.23 (SEQID NO:177), DP II-22.24 (SEQ ID NO: 178), DP II-22.25 (SEQ ID NO:179),or DP II-22.14 (SEQ ID NO:180); and D is DP II-25.2 (SEQ ID NO:71), DPII-25.3 (SEQ ID NO:148), DP II-25.4 (SEQ ID NO: 149), DF II-25.11 (SEQID NO:182), DP II-25.9 (SEQ ID NO:183), DF II-25.10 (SEQ ID NO:184), orDF II-25.13 (SEQ ID NO:186), DP II-25-14 (SEQ ID NO:187), DP II-25.15(SEQ ID NO:188), DP II-25.16 (SEQ ID NO:189), DP II-25.17 (SEQ IDNO:190), DP II-25.18 (SEQ ID NO:191); with the proviso that X, Y, Z, A,B, C, D is not the following combination of peptides: DP I-21.2 (SEQ IDNO:91), DF I-22.2 (SEQ ID NO: 93), DP I-23.1 (SEQ ID NO: 95), DP I-26.1(SEQ ID NO: 99), DP II-20.6, (SEQ ID NO: 56) DP II-22 (SEQ ID NO: 63),and DP II-25.2 (SEQ ID NO: 71).

The present invention also provides novel modified Der p I, Der f I, andDer p II peptides which are a part of a preformulation scheme to developan optimized drug product for therapeutic treatment of humans sufferingfrom allergy to house dust mite allergen. Such peptides and modifiedpeptides possess certain unique characteristics which render themparticularly suitable for drug product formulation, and may be referredto herein as "unique" peptides.

In accordance with pharmaceutical chemistry, preformulation is theprocess of optimizing a drug through determination and/or definition ofthose physical and chemical properties considered important in theformulation of a stable, effective, and safe dosage form. The possibleinteractions with the various components intended for use in the finaldrug product are also considered. Preformulation is an intensive effortthat includes the study of such parameters as solubility, pH profile ofstability, and drug-excipient interactions, which may have a profoundeffect on a drug's physiological availability and physical and chemicalstability. The data obtained from such studies are integrated with thoseobtained from preliminary pharmacological and biochemical studies of theactive drug component thus providing information that permits theselection of the best drug form, and the most desirable excipients foruse in its development.

The development of an optimum formulation of active drug component andexcipients is complex and many factors influence formulation properties.The high degree of uniformity, the physiological availability and thetherapeutic quality expected of pharmaceuticals can only be achieved byconsiderable effort and expertise. Flexibility is also an importantfactor in preformulation. Numerous excipients, stabilizers counter ionsand the like may have to be tested in order to find those compatiblewith the active drug component of the formulation. Multiplemodifications of the active component may become necessary tosuccessfully formulate a drug product. Such modifications must noteffect the overall therapeutic effectiveness of the drug but at the sametime, must render the drug more suitable for formulation.

As a part of a preformulation scheme to provide an optimized drugproduct suitable for use in humans and other mammals for treatingsensitivity to house dust mite, it was determined that the activecomponent (referred to herein as a "peptide" or "candidate peptide" or"unique peptide") in such formulation should possess the followingcharacteristics which would render such peptides "unique" among all ofthe possible peptides derived from the Der p I, Der p II and Der f Iprotein allergen sequences. First, a unique peptide should alone or incombination with other unique peptides comprise a sufficient percentageof the T cell reactivity of the Der p and Der f protein allergens toinduce T cell nonresponsiveness or reduced T cell responsiveness in asubstantial percentage of the individuals sensitive to house dust miteallergen. Second, the candidate peptide should possess thecharacteristic of "superior solubility" which is defined herein assolubility of greater than 3 mg/mi at a pH in a pH range of pH 6 to pH 8in an aqueous buffer. Third, the peptide is stable in an aqueous bufferat a pH in a pH range from pH 6 to pH 8. Candidate peptides derived fromDer p and Der f protein allergens which have been determined to be"unique" peptides of the invention are DPI-21.2 (SEQ. ID. NO. 28),DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165), DFI-26.6 (SEQ.ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14 (SEQ. ID. NO.180) and DPII-25.15 (SEQ. ID. NO. 188) all as shown in FIG. 33.

In accordance with the first characteristic, Those peptides found toelicit a T cell response such as T cell proliferation or lymphokinesecretion (i.e. comprise at least one T cell epitope), or induce T cellnon-responsiveness or reduced T cell responsiveness are understood tohave T cell reactivity. It is believed that exposure of house dust miteallergic patients to isolated house dust mite Group I and Group IIprotein allergen peptides which comprise at least one T cell epitope maycause T cell non-responsiveness of appropriate T cell subpopulationssuch that they become unresponsive or have reduced responsiveness to theprotein allergen and do not participate in stimulating an immuneresponse upon such exposure for example, via anergy, tolerance, orapoptosis, the ability to modify the lymphokine secretion profile ascompared with exposure to the naturally occurring autoantigen; and/orthe ability to cause induction of T suppresser cells.

As discussed earlier, to determine peptides having T cell reactivity andcomprising at least one T cell epitope, isolated peptides are tested by,for example, T cell biology techniques, to determine whether thepeptides elicit a T cell response or induce T cell non-responsiveness.As discussed in the Examples human T cell stimulating activity can betested by culturing T cells obtained from an individual sensitive tohouse dust mite allergen, (i.e., an individual who has an IgE mediatedimmune response to house dust mite allergen) with a peptide or modifiedpeptide derived from a Der p or Der f Group I or Group II proteinallergen and determining whether proliferation of T cells occurs inresponse to the peptide as measured, e.g., by cellular uptake oftritiated thymidine. Stimulation indices for responses by T cells topeptides can be calculated as the maximum counts per minute (CPM) inresponse to a peptide divided by the control CPM. A stimulation index(S.I.) equal to or greater than two times the background level isconsidered "positive". Positive results are used to calculate the meanstimulation index for each peptide for the group of patients tested.Peptides suitable as candidates for formulation into a final drugproduct have a mean T cell stimulation index of greater than or equal to2.0 and preferably higher, (e.g. at least 2.5, more preferably at least3.5, more preferably at least 4.0, more preferably at least 5, even morepreferably at least 7 and most preferably at least about 9).

For therapeutic purposes, candidate peptides are recognized by at least10%, more preferably at least 20%, more preferably at least 30% and evenmore preferably at least 40% or more of individuals in a population ofindividuals sensitive to house dust mite allergen. In addition,preferred candidate peptides have a positivity index (P.I.) of at leastabout 100, more preferably at least about 250 and most preferably atleast about 350. The positivity index for a peptide is determined bymultiplying the mean T cell stimulation index by the percent ofindividuals, in a population of individuals sensitive to house dust miteallergen (e.g., preferably at least 15 individuals, more preferably atleast 30 individuals or more), who have a T cell stimulation index tosuch peptide of at least 2.0. Thus, the positivity index represents boththe strength of a T cell response to a peptide (S.I.) and the frequencyof a T cell response to a peptide in a population of individualssensitive to house dust mite allergen.

To determine whether a peptide (candidate peptide) or a combination ofcandidate peptides are likely to comprise a sufficient percentage of theT cell reactivity of house dust mite protein allergens, to induce T cellnonresponsiveness in a substantial percentage of a population ofindividuals sensitive to house dust mite allergen, an algorithm can beused. In accordance with one such algorithm, a set of overlappingpeptides is produced by systematically dividing the protein allergen(s)of interest into at least two overlapping peptide regions of desiredlengths (e.g., of about 12-30 amino acid residues in length, preferablynot longer than about 25 amino acid residues in length with about 5-15amino acid residues of overlap). This division into peptide regions canbe arbitrary, can be made according to an algorithm, or can be wholly orpartially based on regions of house dust mite Group I and/or Group IIprotein allergens known to comprise at least one T cell epitope.Preferably, at least 50% of the entire house dust mite protein allergensequence and more preferably, the entire house dust mite proteinallergen sequence is divided into two or more peptides. A human T cellstimulation index is determined for each of the peptides in an in vitroT cell proliferation assay as described herein for each individualtested in a population of individuals sensitive to the protein antigen.For example both WO93/08279 and WO94/24281 disclose T cell studies withoverlapping peptides derived from Der p I, Der p II, Der f I and Der fII. A candidate peptide or combination of candidate peptides is selectedbased, at least in part, on the mean human T cell stimulation index ofthe candidate peptide in the set of peptides tested and the positivityindex of the candidate peptide in the set of peptides tested (see, FIGS.3 and 4). The human T cell stimulation index for the candidatepeptide(s) is summed. For each individual, the human T cell stimulationindex for the candidate peptide(s) is divided by the sum of the human Tcells stimulation indices of the remaining peptides in the set ofpeptides tested to determine a percent of T cell reactivity as shownbelow: ##EQU1##

Alternatively, the presence of T cell epitopes in the candidate peptidedependent on amino acids residues in an overlapping peptide located ateither the N-terminus or C-terminus of the candidate peptide in theamino acid sequence of the protein antigen, but which epitopes are notpresent in the candidate peptide can be considered in calculating thepercent of T cell reactivity in the candidate peptide by use of thefollowing formula: ##EQU2##

In this formula, "N_(T) flanking peptide" refers to a peptide whichcomprises amino acid residues which overlap with amino acid residueslocated at the N-terminus of the candidate peptide in the amino acidsequence of the protein antigen from which the peptide is derived;"C_(T) flanking peptide" refers to a peptide which comprises amino acidresidues which overlap with amino acid residues located a the C-terminusof the candidate peptide in the amino acid sequence of the proteinantigen from which the peptide is derived. In this calculationstimulation indices for the candidate peptide, the N-terminal flankingpeptide and the C-terminal flanking peptide are added and divided by thesum total of the stimulation indices for the entire set of overlappingpeptides obtain a percent of T cell reactivity for the candidatepeptide. If a combination of two or more candidate peptides is selectedeach of which contains amino acid residues which overlap, thiscalculation cannot be used to determine a percent of T cell reactivityfor each candidate peptide separately. However, a total percent of Tcell reactivity for the combination of candidate peptides can beobtained. In this situation, the stimulation indices for all of thecandidate peptides which overlap is included in the calculation.

The values obtained for the percentage of T cell reactivity for thecandidate peptide or combination of peptides in each individual testedcan be expressed as a range of the lower and higher values of theresults of the above described calculations. By either of the abovecalculations, the percent is obtained for at least about twenty (20) andpreferably at least about thirty (30) individuals sensitive to theprotein antigen and a mean percent is determined. For use in thecompositions of the invention, the candidate peptide or combination ofcandidate peptides has the following criteria: (1) the candidate peptideor combination of candidate peptides has a mean percent of at leastabout 10%, preferably at least about 20%, more preferably at least about30%, more preferably at least about 40% and more preferably at leastabout 50-60% or greater; and (2) in the population of individuals testedat least about 60%, preferably at least about 75%, and more preferablyat least about 90-100% have positive T call responses (S.I. equal to orgreater than 2.0) in response to the candidate peptide or combination ofcandidate peptides. A candidate peptide or combination of candidatepeptides meeting the above criteria is likely to comprise a sufficientpercentage of the T cell reactivity to house dust mite protein allergento induce T cell non-responsiveness or reduced T cell responsiveness ina substantial percentage of a population of individuals sensitive tohouse dust mite.

As an illustrative embodiment of the above-described algorithm, a set ofoverlapping peptides and candidate peptides derived from Der p I and Derp II respectfully were produced and tested. Secondary T cell culturesdetermined to be reactive with Der p I protein allergen were derivedfrom 39 house dust mite-allergic subjects and analyzed for reactivity toan overlapping set of peptides, as well as candidate peptides derivedfrom Der p I and Der f I protein allergen, DPI-21.2 (SEQ. ID. NO. 28),DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165) and DFI-26.6(SEQ. ID. NO. 168), in an in vitro T cell proliferation assay asdescribed herein. The results are shown in FIG. 3. The higheststimulation index greater than or equal to 2.0 in response to eachpeptide was recorded for each subject tested. The data were thenanalyzed by the equations above. The results and calculations of thepercent of T cell reactivity for a single dust mite-allergic subject areshown below using formulas (1) and (2).

T CELL REACTIVITY FOR PATIENT 1733

    ______________________________________                                        PEPTIDE STIMULATION INDEX                                                     ______________________________________                                        DPI-21.2 (SEQ. ID. NO. 28)                                                                         3.6                                                      DPI-3 (SEQ. ID. NO. 11)                                                                            3.9                                                      DPI-22.2 (SEQ. ID. NO. 30)                                                                         3.1                                                      DPI-12.1 (SEQ. ID. NO. 14)                                                                         2.2                                                      DPI-5.1 (SEQ. ID. NO. 205)                                                                         3.5                                                      DFI-23.31 (SEQ. ID. NO. 165)                                                                       5.7                                                      DPI-14 (SEQ. ID.NO. 18)                                                                            2.3                                                      DPI-15 (SEQ. ID. NO.)                                                                              2.8                                                      DPI-6.1 (SEQ. ID. NO. 19)                                                                          2.1                                                      DPI-7.1 (SEQ. ID. NO. 21)                                                                          2.2                                                      DFI-26.6 (SEQ. ID. NO. 168)                                                                        5.0                                                      DPI-9 (SEQ. ID. NO. 23)                                                                            2.4                                                      DPI-16 (SEQ. ID. NO. 24)                                                                           2.0                                                      DPI-10 (SEQ. ID. NO. 25)                                                                           0                                                        DPI-17 (SEQ. ID. NO. 26)                                                                           0                                                        SUM OF STIMULATION INDICES                                                                        40.8 (DENOMINATOR)                                        ______________________________________                                         % Reactivity of Peptide DFI26.6 (SEQ. ID. NO. 168) for patient 1733 is        ##STR1##                                                                      ##STR2##                                                                 

Therefore the estimated range of T cell reactivity for Peptide DFI-26.6(SEQ. ID. NO. 168) for this patient is 12.3%-24% of the total reactivityof the Der p I protein. The above calculation is repeated for anypotential candidate peptides for each patient tested. In the populationof 39 Cry j I-allergic subjects tested the following results wereobtained:______________________________________ Range ofCandidate meanpercentage Frequency of responsePeptides T Cell Reactivity at least onepeptide______________________________________DPI-21.2(SEQ. ID. NO.28),DFI-22.2(SEQ. ID. NO. 93)DPI-23.31 38-67% 82%(SEQ. ID. NO.165),DPI-26.6(SEQ. ID. NO. 168)______________________________________

Thus, the combination of the four candidate peptides are well within thedesired range for possessing, in combination, sufficient T cellreactivity of Group I protein allergen of Der f and Der p, and meet thefirst characteristic of a "unique" peptide of the invention.

The same calculations were determined for the Group II, Der p proteinallergen. Secondary T cell cultures determined to be reactive with Der pI protein allergen were derived from 30 house dust mite-allergicsubjects and analyzed for reactivity to an overlapping set of peptides,as well as candidate peptides derived from Der p II, DPII-20.9 (SEQ. ID.NO. 169), DPII-22.14 (SEQ. ID. NO. 180)and DPII-25.15 (SEQ. ID. NO.188), in an in vitro T cell proliferation assay as described herein. Theresults are shown in FIG.4. The highest stimulation index greater thanor equal to 2.0 in response to each peptide was recorded for eachsubject tested. The data were then analyzed by the equations above. Inthe population of 30 house dust mite-allergic subjects tested thefollowing results were obtained:______________________________________Range ofCandidate mean percentage Frequency of responsePeptides T CellReactivity at least onepeptide______________________________________DPII-20.9(SEQ. ID. NO.169),DPII-22.14(SEQ. ID. NO. 180)DPII-25.15 37-51% 63%(SEQ. ID. NO.188)______________________________________

Thus, the combination of the three Der p II candidate peptides of theinvention are well within the desired range for possessing, incombination, sufficient T cell reactivity of Group II house dust miteprotein allergen of Der p, and meet the first characteristic of a"unique" peptide of the invention.

For the treatment of allergy in accordance with the methods of theinvention, it is preferred that a peptide used in conjunction therewithdoes not bind immunoglobulin E (IgE) or binds IgE to a substantiallylesser extent (i.e. at least 100-fold less binding and more preferably,at least 1,000-fold less binding) than the respective house dust miteprotein allergen from which the peptide is derived binds IgE.Experiments to date indicate that candidate peptides DPI-21.2 (SEQ. ID.NO. 28), DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165),DFI-26.6 (SEQ. ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14(SEQ. ID. NO. 180) and DPII-25.15 (SEQ. ID. NO. 188) exhibit negativeIgE reactivity or histamine release (data not shown).

The second characteristic for a unique peptide is that of "superiorsolubility" which was defined earlier as being solubility of greaterthan 3 mg/ml at a pH in a range of pH 6 to pH 8. Solubility in aphysiologically acceptable pH range (e.g. pH 6 to pH 8) is particularlyimportant when formulating a multipeptide therapeutic for injection.Administration of a soluble drug product in a physiologically acceptablepH range by intravenous or subcutaneous injection provides about 100%bioavailability of the drug component to the physiological system intowhich the drug is being introduced. Thus, it is necessary that a drugproduct intended for injection be fluid to the extent that easysyringability exists, and the active component be soluble as well ifmaximum therapeutic effect is to be achieved. Solubility is also usefulwhen formulating compositions to be administered via other modes ofadministration such as by oral administration (tablet, aerosol,sublingual), or sustained release preparations and formulations.

Proteins and peptides may be difficult to formulate into solublecompositions as a peptide may not be soluble in any desirable pH rangeor may be soluble in only a narrow pH range. It is particularlydifficult when multiple peptides are being formulated together into asingle multipeptide formulation, as each peptide may be soluble in a pHrange which does not overlap with those of the other peptides in theformulation. As a result, it is the requirement of "superior solubility"which requires the most formulation flexibility in that considerablemodification of the targeted candidate peptides may be necessary tosuccessfully formulate a multipeptide drug product.

Some of the unique peptides of the invention are the product of multipleamino acid modifications of the original targeted candidate peptidesequence ("parent") from which the modified unique peptides of theinvention were originally derived. Such modified "unique" peptides ofthe invention include DFI-23.31 (SEQ. ID. NO. 165), DFI-26.6 (SEQ. ID.NO. 168), DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14 (SEQ. ID. NO. 180)and DPII-25.15 (SEQ. ID. NO. 188) all as shown in FIG. 33. For example,the amino acid sequence of DPII-22.14 (SEQ. ID. NO. 180) was derivedfrom the protein sequence of Der p II by first identifying those regionsof the native protein with high T-cell reactivity using a set ofoverlapping peptides that covered the entire protein sequence. Severalpeptides were found to exhibit high T-cell reactivity, corresponding tothree different locations/areas in the protein. One of these areas werecovered by three adjacent peptides each being about 25-29 amino acidslong and with 10-15 amino acids overlap, DpII-3.1 (SEQ. ID. NO. 206),DpII-4 (SEQ. ID. NO. 44)and DpII-5 (SEQ. ID. NO. 45) (FIG. 34) that allexhibited high T-cell reactivity. Based on this T-cell map a new peptidewas synthesized, with the amino acid sequenceQLEAVFEANQNTKTAKIEIKASIDGLEV (SEQ. ID. NO. 207), 28 amino acids long,which exhibits most of the reactivity found in the individual peptides(data not shown) and contains all of DpII-4 (SEQ. ID. NO. 44)and partsof DpII-3.1 (SEQ. ID. NO. 206) and DpII-5 (SEQ. ID. NO. 45). However,this peptide did not meet the "superior solubility" requirement of 3mg/ml for a "unique" peptide of the invention nor did it meet thestability requirement. Therefore two analogs were made with addition ofone or two charged residues at each end, DpII-22.2KQLEAVFEANQNTKTAKIEIKASIDGLEVK (SEQ. ID. NO. 93) and DpII-22.6DKQLEAVFEANQNTKTAKIEIKASIDGLEVD (SEQ. ID. NO. 139) and the stabilityimproved substantially but the solubility did not reach the standard ofa "superior solubility". In the second attempt a series of truncatedanalogs with up to five charged amino acid residues added to the ends(Table 1).

                                      TABLE 1                                     __________________________________________________________________________    DpII-22 (SEQ. ID. NO. 63)                                                                    QLEAVFEANQNTKTAKIEIKASIDGLEV                                   DpII-22.2 (SEQ. ID. NO. 93)                                                                  KQLEAVFEANQNTKTAKIEIKASIDGLEVK                                 DpII-22.6 (SEQ. ID. NO. 139)                                                                 DKQLEAVFEANQNTKTAKIEIKASIDGLEVD                                DpII-22.4 (SEQ. ID. NO. 141)                                                                 QLEAVFEANQNTKTAKIEIKASIDE                                      DpII-22.5 (SEQ. ID. NO. 142)                                                                 DKQLEAVFEANQNTKTAKIEIKASIDE                                    DpII-22.8 (SEQ. ID. NO. 144)                                                                 DKEQLEAVFEANQNTKTAKIEIKASIDE                                   DpII-22.9 (SEQ. ID. NO. 145)                                                                 DKEQLEAVFEANQNTKTAKIEIKASIDEE                                  DpII-22.12 (SEQ. ID. NO. 198)                                                                DKQLEAVFEANQATKTAKIEIKASIDE                                    DpII-22.16 (SEQ. ID. NO. 199)                                                                DKELEAVFEANQNTKTAKIEIKASD                                      DpII-22.19 (SEQ. ID. NO. 200)                                                                DKELEAVFEANQNTKTAKIEIKAD                                       DpII-22.20 (SEQ. ID. NO. 201)                                                                DKELEAVFEANQNTKTAKIEIKAK                                       DpII-22.21 (SEQ. ID. NO. 202)                                                                DKELEAVFEANQNTKTAKIEIKD                                        DpII-22.22 (SEQ. ID. NO. 175)                                                                DKELEAVFEANQNTKTAKIEIKK                                        DPII-22.26 (SEQ. ID. NO. 176)                                                                DKELEAVFEANQNTKTAKIEIK                                         DpII-22.23 (SEQ. ID. NO. 177)                                                                DKELEAVFEANQNTKTAKIED                                          DpII-22.24 (SEQ. ID. NO. 178)                                                                DKELEAVFEANQNTKTAKIEK                                          DpII-22.25 (SEQ. ID. NO. 179)                                                                DKELEAVFEANQNTKTAKIE                                           DPII-22.14 (SEQ. ID. NO. 180)                                                                DKELEAVFEANQNTKTAKAE                                           DpII-22.10 (SEQ. ID. NO. 146)                                                                LEAVFEANQNTKTAK                                                DpII-22.11 (SEQ. ID. NO. 147)                                                                LEAVFEANQATKTAK                                                DpII-22.18 (SEQ. ID. NO. 203)                                                                DKTAKIEIKASIDGLE                                               DpII-22.15 (SEQ. ID. NO. 204)                                                                KTAKIEIKASIDGLE                                                __________________________________________________________________________

Of these the DpII-22.5, DKQLEAVFEANQNTKTAKIEIKASIDE (SEQ. ID. NO. 142),was promising since it retained the T-cell reactivity of the "parent"peptide, DPII-22, (SEQ. ID. NO. 63) and was very soluble, but it wasvery difficult to synthesize. The difficulties in synthesizing thissequence were found to disappear with the replacement of the hydrophobicisoleucine with the less hydrophobic alanine and simultaneously thesolubility increased by an order of magnitude. Peptide DpII-22.14,DKELEAVFEANQNTKTAKAE (SEQ. ID. NO. 180), was found to possess almost thesame T-cell reactivity as the "parent" peptide DpII-22 (SEQ. ID. NO.63)as well as being soluble at greater than 3 mg/ml at a pH in the pHrange 6.0 to 8.0, and was easy to synthesize and purify. Therefore,DPII-22 (SEQ. ID. NO. 63) was chosen as a "unique" peptide when it wasdetermined to be stable at a pH in the range of 6.0 to 8.0. Thedevelopment of the other modified "unique" peptides, DFI-23.31(SEQ. ID.NO. 165), DFI-26.6 (SEQ. ID. NO. 168) and DPII-25.15 (SEQ. ID. NO. 188),followed a process similar to that described above for DPII-22.14 (SEQ.ID. NO. 180). All of the unique candidates are novel peptides of thisinvention.

The third criteria which the unique peptides of this invention must meetis stability, particularly solution stability, in a physiologicallyacceptable pH range of pH 6 to pH 8. It must be stable under theconditions of manufacture and storage, and under conditions ofreconstitution if necessary. Stability testing establishes the timeperiod for which the integrity, quality and purity of the drug productis preserved in its finished dosage form. Stability testing may beperformed concurrently with solubility studies as discussed in Example3. An equal concentration composition comprising each of the candidatepeptides of the invention remained stable (e.g. no significantdegradation) in solution at a common "window" within the pH range frompH 6-pH 8 at about room temperature and at about 5° C. for at least 24hours (see, FIG. 6).

Therefore, candidate peptides DPI-21.2 (SEQ. ID. NO. 28), DFI-22.2 (SEQ.ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165), DFI-26.6 (SEQ. ID. NO. 168),DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14 (SEQ. ID. NO. 180) andDPII-25.15 (SEQ. ID. NO. 188) possess each of the three required"unique" characteristics outlined above, indicating that thiscombination of peptides is suitable for formulation as an optimizedtherapeutic drug product for administration to humans for treatment ofallergy to house dust mite allergen.

Highly purified peptides of this invention, may be producedsynthetically by chemical synthesis using standard techniques. Variousmethods of chemically synthesizing peptides are known in the art such assolid phase synthesis which has been fully or semi automated oncommercially available peptide synthesizers. Synthetically producedpeptides may then be purified to homogeneity (i.e. at least 90%, morepreferably at least 95% and even more preferably at least 97% purity),free from all other polypeptides and contaminants using any number oftechniques known in the literature for protein purification.

In accordance with one procedure for producing highly purifiedhomogenous peptides of the invention, a peptide produced by syntheticchemical means (either anchored to a polymer support "solid phasesynthesis" or by conventional homogenous chemical reactions "solutionsynthesis") may be purified by preparative reverse phase chromatography.In this method, the synthetically produced peptide in "crude" form isdissolved in an appropriate solvent (typically an aqueous buffer) andapplied to a separation column (typically a reverse phase silica basedmedia, in addition, polymer or carbon based media may be used). Peptideis eluted from the column by increasing the concentration of an organiccomponent (typically acetonitrile or methanol) in an aqueous buffer(typically TFA, triethylamine phosphate, acetate or similar buffer).Fractions of the eluate will be collected and analyzed by appropriateanalytical methods (typically reverse phase HPLC or CZE chromatography).Those fractions having the required homogeneity will be pooled. Thecounter ion present may be changed by additional reverse phasechromatography in the salt of choice or by ion exchange resins. Thepeptide may then be isolated as its acetate or other appropriate salt.The peptide is then filtered and the water removed (typically bylyophilization) to give a homogenous peptide composition containing atleast 90%, more preferably at least 95% and even more preferably atleast 97% of the required peptide component. Optionally, or inconjunction with reverse phase HPLC as described above, purification maybe accomplished by affinity chromatography, ion exchange, sizeexclusion, counter current or normal phase separation systems, or anycombination of these methods. Peptide may additionally be concentratedusing ultra filtration, rotary evaporation, precipitation, dialysis orother similar techniques.

The highly purified homogenous peptide composition is then characterizedby any of the following techniques or combinations thereof: a) massspectroscopy to determine molecular weight to check peptide identity; b)amino acid analysis to check the identity of the peptide via amino acidcomposition; c) amino acid sequencing (using an automated proteinsequencer or manually) to confirm the defined sequence of amino acidresidues; d) HPLC (multiple systems if desired) used to check peptideidentity and purity (i.e. identifies peptide impurities); e) watercontent to determine the water concentration of the peptidecompositions; f) ion content to determine the presence of salts in thepeptide composition; and g) residual organics to check for the presenceof residual organic reagents, starting materials, and/or organiccontaminants.

A peptide of the invention may also be produced by recombinant DNAtechniques in a host cell transformed with a nucleic acid sequencecoding for such peptide as described earlier.

In certain limited circumstances, peptides of this invention may also beproduced by chemical or enzymatic cleavage of a highly purified fulllength or native protein of which the sites of chemical digest orenzymatic cleavage have been predetermined and the resulting digest isreproducible. Peptides having defined amino acid sequences can be highlypurified and isolated free of any other poly peptides or contaminantspresent in the enzymatic or chemical digest by any of the proceduresdescribed above for highly purified, and isolated synthetically orrecombinantly produced peptides.

The present invention also pertains to therapeutic compositions andmultipeptide therapeutic formulations comprising the unique peptides ofthe invention. Therapeutic compositions of the invention may compriseone or more of the unique peptides of the invention which may beadministered simultaneously or sequentially as single treatment episodefor treatment of allergy to house dust mite allergen in a human or othermammal. Such a treatment regimen may not necessarily be a physicalmixture of more than one peptide of the invention, but does comprise acombination of such peptides administered simultaneously or sequentiallyas a single treatment episode in order to achieve the maximumtherapeutic effect the combination of the unique peptides, DPI-21.2(SEQ. ID. NO. 28), DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO.165), DFI-26.6 (SEQ. ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169),DPII-22.14 (SEQ. ID. NO. 180) and DPII-25.15 (SEQ. ID. NO. 188) provide(e.g. solubility and stability in acceptable physiological pH range aswell as having a range of T cell reactivity of 38-67% of the total Tcell reactivity of the Group I house dust mite protein allergen and afrequency of response of 91% for at least one Der p I peptide tested ina population of individuals allergic to house dust mite and similarly,have a range of T cell reactivity of 37-51% of the total T cellreactivity of the Group II house dust mite protein allergen and afrequency of response of 63% for at least one Der p II peptide tested ina population of individuals allergic to house dust mite).

Therapeutic compositions of the invention comprise one or more ofpeptides DPI-21.2 (SEQ. ID. NO. 28), DFI-22.2 (SEQ. ID. NO. 93),DPI-23.31(SEQ. ID. NO. 29), DFI-26.6 (SEQ. ID. NO. 168), DPII-20.9 (SEQ.ID. NO. 169), DPII-22.14 (SEQ. ID. NO. 180) and DPII-25.15 (SEQ. ID. NO.188) also comprise one or more pharmaceutically acceptable carriers suchas excipients which are compatible with peptide or peptides present in asingle composition. When the composition is a multipeptide formulation,the pharmaceutically acceptable carrier must be compatible with all ofthe peptides in the multipeptide formulation. Preferred excipientsinclude but are not limited to sterile water, sodium phosphate,mannitol, or both sodium phosphate and mannitol or any combinationthereof. Other suitable excipients include but are not limited tosorbitol, sucrose, dextrose, lactose dextran and PVP. Additionally, dueto the potential for dimerization of the peptides in a mutlipeptideformulation, there may also be included an agent such as EDTA to preventdimerization. Alternatively, any other material or procedures known inthe art to prevent dimerization may be used. In addition,pharmaceutically acceptable counter ions may be added during thepreparation of the multipeptide formulation. Examples ofpharmaceutically acceptable counter ions include acetate, HCl, andcitrate.

A therapeutic composition of the invention should be sterile, stableunder conditions of manufacture, storage, distribution and use andshould be preserved against the contaminating action of microorganismssuch as bacteria and fungi. A preferred means for manufacturing atherapeutic compositions of the invention in order to maintain theintegrity of the composition (i.e. prevent contamination, prolongstorage, etc.) is to prepare the formulation of peptide andpharmaceutically acceptable carrier(s) such that the composition may bein the form of a lyophilized powder which is reconstituted in apharmaceutically acceptable carrier, such as sterile water, just priorto use. In the case of sterile powders for the preparation of sterileinjectable solutions, the preferred methods of preparation are vacuumdrying, freeze-drying or spin drying which yields a powder of the activeingredient plus any additional desired ingredient from a previouslysterile-filtered solution thereof.

A preferred multipeptide formulation comprises the following uniquepeptides DPI-21.2 (SEQ. ID. NO. 28), DFI-22.2 (SEQ. ID. NO. 93),DFI-23.31(SEQ. ID. NO. 165), DFI-26.6 (SEQ. ID. NO. 168), DPII-22.14(SEQ. ID. NO. 180) and DPII-25.15 (SEQ. ID. NO. 188) and sodiumphosphate and mannitol, and optionally may further comprise DPII-20.9(SEQ. ID. NO. 169). For this embodiment, EDTA is added to theformulation. A suitable counter ion such as acetate may also be addedduring the preparation of the formulation. The formulation is preferablyprepared in the form of a lyophilized powder which is reconstituted in aphysiologically acceptable carrier, such as sterile water, prior to use.Several non-limiting examples of a preferred multipeptide formulationsof this invention are described below. The unique house dust miteprotein allergen peptides will preferably be combined duringmanufacturing with the appropriate counter ion to produce one vialcontaining a sterile, pyrogen free, and preferably lyophilized powder ofthe desired formulation:

Active: Der p I, Der f I and Der p II peptides DPI-21.2 (SEQ. ID. NO.27), DFI-22.2 (SEQ. ID. NO. 93), DPI-23.31 (SEQ. ID. NO. 29), DFI-26.6(SEQ. ID. NO. 168), DPII-22.14 (SEQ. ID. NO. 32) and DPII-25.15 (SEQ.ID. NO. 188)

In concentration of 0.75 mg per peptide

Inactives: 0.05M Sodium Phosphate U.S.P.

5% w/v Mannitol, U.S.P.

0.1 mg/ml EDTA disodium dihydrate U.S.P.

Final pH 7.2-7.4

Diluent: Sterile Water for Injection, U.S.P.

Optionally 0.75 mg of DPII-20.9 (SEQ. ID. NO. 169) may be added to theactive ingredients.

A preferred combination of multipeptide formulations suitable foradministration simultaneously or sequentially as a single treatmentepisode and contained in two separate sterile, pyrogen free vialspreferably in the form of lyophilized powders include the followingformulations:

Vial #1

Active: Der p I, Der f I and Der p II peptides DFI-22.2(SEQ. ID. NO.28), DFI-23.31(SEQ. ID. NO. 165), and DPII-22.14 (SEQ. ID. NO. 32)

In concentration of 0.75 mg per peptide

Inactives: 0.05M Sodium Phosphate U.S.P.

5% w/v Mannitol, U.S.P.

0.1 mg/ml EDTA disodium dihydrate

Final pH 7.0

Diluent: Sterile Water for Injection, U.S.P.

Vial #2

Active: Der p I, and Der p II peptides DPI-21.2 (SEQ. ID. NO. 28),DFI-26.6 (SEQ. ID. NO. 168), DPII 20.9 (SEQ. ID. NO. 31), DPII-25.15(SEQ. ID. NO. 188)

In concentration of 0.75 mg per peptide

Inactives: 0.05M Sodium Phosphate U.S.P.

5% w/v Mannitol, U.S.P.

0.1 mg/ml EDTA disodium dihydrate U.S.P.

Final pH 6.2

Diluent: Sterile Water for Injection, U.S.P.

The multipeptide formulations of the invention may also be provided inthe form of a kit, including instructions for use.

Administration of the therapeutic compositions and multipeptideformulations described above to an individual, preferably innon-immunogenic form, can be carried out using known procedures atdosages and for periods of time effective to cause down regulation ofthe house dust mite-specific immune response (i.e., reduce the allergicsymptoms caused by house dust mite protein allergen of the individual.Down regulation of the allergic immune response to house dust miteallergen in humans may be determined clinically whenever possible (seee.g., Varney et al, British Medical Journal, 302:265-269 (1990), or maybe determined subjectively (i.e. the patient feels as if some or all ofthe allergic symptoms caused by house dust mite allergens have beenalleviated).

One of the unique characteristics of each unique peptide in accordancewith the invention is that each peptide possesses "superior solubility".Therefore, compositions and multipeptide formulations of the inventionare particularly suitable for administration by injection (e.g.subcutaneous, or intravenous). However, optimized compositions andmultipeptide formulations of the invention may be administered in anyconvenient manner wherein solubility of the active drug component iseither desirable or acceptable, such as by injection (subcutaneous,intravenous, etc.), oral administration, sublingual, inhalation,transdermal application, rectal administration, or any other route ofadministration known in the art for administering soluble therapeuticagents. It may be desirable to administer simultaneously or sequentiallya therapeutically effective amount of one or more of the therapeuticcompositions of the invention to an individual as a single treatmentepisode. Each of such compositions for administration simultaneously orsequentially as a single treatment episode, may comprise only one uniquepeptide of the invention or may comprise an optimized multipeptideformulation in accordance with the invention.

For subcutaneous injection of one or more therapeutic compositions andmultipeptide formulations of the invention, preferably about 1 mg-3 mgand more preferably from about 20 mg-1.5 mg, and even more preferablyabout 50 mg-750 mg of each active component (peptide) per dosage unitmay be administered. It is especially advantageous to formulateparenteral compositions in unit dosage form for ease of administrationand uniformity of dosage. Unit dosage form as used herein refers tophysically discrete units suited as unitary dosages for human subjectsto be treated; each unit containing a predetermined quantity of activecompound calculated to produce the desired therapeutic effect inassociation with the desired pharmaceutical carrier. The specificationfor the novel unit dosage forms of the invention are dictated by anddirectly dependent on (a) the unique characteristics of the activecompound and the particular therapeutic effect to be achieved, and (b)the limitations inherent in the art of compounding such an activecompound for the treatment of human subjects.

Effective amounts of the optimized drug compositions of the inventionwill vary according to factors such as the degree of sensitivity of theindividual to the antigen, the age, sex, and weight of the individual,and the ability of peptide to cause down regulation of the antigenspecific immune response in the individual. Dosage regimen may beadjusted to provide the optimum therapeutic response. For example,several divided doses may be administered over the course of days,weeks, months or years, or the dose may be proportionally increased orreduced with each subsequent injection as indicated by the exigencies ofthe therapeutic situation. In one preferred therapeutic regimen,subcutaneous injections of therapeutic compositions are given once aweek for 3-6 weeks. The dosage may remain constant for each injection ormay increase or decrease with each subsequent injection. A boosterinjection may be administered at intervals of about three months toabout one year after initial treatment and may involve only a singleinjection or may involve another series of injections similar to that ofthe initial treatment.

This invention is further illustrated by the following non-limitingexamples.

EXAMPLE I Native Mite Allergen Purification

What follows is a description of the work done to purify the group I andgroup II allergens of the house dust mites Dermatophagoidespteronyssinus and Dermatophagoides farinae in their native form asprimary antigens for human T-cell epitope mapping.

All four protein allergens, Der f I, Der p I, Der f II, and Der p II,were immunoaffinity purified from spent mite culture media obtained fromeither Commonwealth Sera Laboratories Melbourne, Australia or Dr. LarryG. Arlian at Wright State University Dayton, Ohio. A 10% (wt./vol.)aqueous extract of dessicated spent mite culture media was prepared inPBS (Phosphate Buffered Saline) with stirring overnight at 4° C.Insoluble material was removed by centrifugation at 10,000×g for 1 hourat 4° C. The supernatant was then filtered on a vacuum manifold throughWhatman #1 paper, and re-centrifuged at 15,000×g for 1 hour at 4° C. Afinal filtration was carried out through a 0.45 m cellulose acetatefilter.

Monoclonal antibodies 4C1 and 6D6 (University of Virginia, N.C.) wereused for immunoaffinity purification of group I or group II miteallergens respectively (Chapman et al., J. Allergy Clin. Immunol.,80:1479-1484 (1987); Heymann et al., J. Allergy Clin. Immunol.,83:1055-1067 (1989)). The 4C1 monoclonal antibody reacts with an epitopeshared by both the Der f I and Der p I proteins; similarly, the 6D6monoclonal antibody reacts with both the Der f II and Der p II proteins.

For each monoclonal antibody, ascites fluid was cut in 50% ammoniumsulfate and the antibodies coupled to CNBr-activated Sepharose 4B(Pharmacia) in 100 mM NaHCO₃, 500 mM NaCl, pH 8.3, overnight at 4° C.

Monoclonal antibody columns were equilibrated in PBS and the filteredextracts loaded at 15 ml per hour. The column was then washed in 20volumes of PBS, after which a more stringent wash of 20 column volumeswas carried out with PBS supplemented with 500 mM NaCl. Proteins wereeluted in 500 mM NaCl, 100 mM glycine pH 11.0, and fractions evaluatedfor protein content by spectrophotometric absorbance at 280 nm. Peakfractions were pooled and dialyzed extensively against PBS, concentratedwith a negative pressure dialysis device (obtained from Amicon, BeverlyMass.) and were used in T-cell epitope mapping studies of the mite groupI and group II allergens. Recovered proteins were obtained at puritiesranging from 80% (group II) to 90% (group I).

Reverse Phase HPLC chromatography was applied to further purify theimmunoaffinity purified group II mite protein allergen according to theconditions in Heymann et al., J. Allergy Clin. Immunol., 83:1055-1067(1989). Briefly, immunoaffinity purified protein was applied to a 5 μm300 Å C-8 column (Applied Biosystems Inc.) in 0.1% vol./vol. TFA/H₂ O.The flow rate for the column was 1 m/min. with a gradient of 0-60%acetonitrile/0. 1% TFA over 60 minutes. Group II proteins eluted around45% acetonitrile/0.1% TFA. Fractions were analyzed for purity bySDS-polyacrylamide gel electrophoresis followed by densitometricscanning. Those fractions with group II protein of purity greater than90% were subsequently utilized for human T-cell mapping of the allergen.

EXAMPLE II Recombinant Mite Allergen Expression

What follows is a description of the work done to produce the group Iand group II allergens of the house dust mites Dermatophagoidespteronyssinus and Dermatophagoides farinae as recombinant proteins in E.coli.

All four protein allergens, Der f I, Der p I, Der f II, and Der p II,were fused at their mature amino termini with the leader sequenceMGHHHHHHEF (SEQ ID NO: 150) (where amino acids EF are encoded by theEcoR I restriction site GAATTC). Since the H₆ stretch of amino acidscoordinates Ni⁺⁺ ions on NTA agarose columns (Diagen GmbH), thisactivity was exploited in the purification of the recombinant proteins(Hochuli et al., Biotechnology, 86:1321-1325 (1988)). Ultimately, allfour allergens were subcloned into the expression vector pET 11d(Studier et al., Methods In Enzymology, 185:60-88 (1990)) under thetranscriptional control of the phage T7 gn 10 promoter.

The cDNAs encoding the group I allergens from D. pteronyssinus and D.farinae were obtained from Dr. Wayne Thomas in plasmid form as subclonesfrom λgt 11 (Chua et al., J. Exp. Med., 167:175-182 (1988); Dilworth etal., Clin. Exp. Allergy, 21: 25-32 (1991)). The Der p I cDNA had beensubcloned from an M13 RF plasmid as an EcoRI fragment into pUC18 by Dr.Roland Buelow at ImmuLogic (Palo Alto), while the Der f I cDNA wasmanipulated directly from the M13mp19 RF plasmid Df I(1).

Initially the cDNAs were subcloned into the expression vector pTrc99AHis₆ Insert RRRS which is a modified version of pTrc99A (Amann et al.,Gene, 69:301-315 (1988)). The original vector was modified by theaddition of a synthetic adaptor (consisting of two complimentaryoligonucleotides) encoding the leader sequence MGHHHHHHEF (SEQ ID NO:150) between its NcoI (MG) and EcoRI (EF) sites at the 5' end of thepolylinker, and an RRS (Retro Regulatory Sequence) into its Hind IIIsite at the polylinker's 3' end. The leader sequence was used as apurification aid as described above, while the RRS was added to enhancerecombinant message stability and thereby increase recombinant proteinyield (Skoglund et al., Gene, 88:1-5 (1990)). Lastly, an insert, in thiscase an Amb a I.1 cDNA encoding the major allergen of the short ragweed(Rafnar et al., J. Biol. Chem., 226:1229-1236 (1991)) was subcloned inframe with the H₆ leader as an EcoRI to PstI fragment.

To express the mite group I allergens, the Amb a I.1 cDNA was excised byEcoRI/Hind III digestion and replaced by adaptors with φEcoRI/Hind IIIoverhangs (composed of a pair of complimentary oligonucleotides). Theseadaptors (FIG. 2a) encoded the first 5 amino acids, up to the Pst Isite, of the mature Der p I NH₂ terminus, and the first 10 amino acids,up to the HpaI site, of the mature Der f I NH₂ terminus. Upon ligationof the EcoRI site of the vector and the φEcoRI site in the adaptors, theEcoRI site in the vector was destroyed, leaving the EcoRI site encodedinternal to the adaptor as the sole EcoRI site in the intermediateconstructs pTrc99A His₆ 5' pl RRS and pTrc99A His₆ 5 ' fl RRS. The Der pI cDNA was inserted as a PstI/EcoRI fragment into PstI/EcoRI digestedpTrc99A His₆ 5' pl RRS, while the Der f I cDNA was inserted as aHpaI/EcoRI fragment into Hpal/EcoRI digested pTrc99A His₆ 5' fl RRS. Thesequence of the 5' end of each construct, pTrc99A His₆ 5' pl RRS andpTrc99A His₆ fl RRS, was verified by dideoxy chain termination DNAsequencing using a Sequenase™ II kit (U.S. Biochemicals). Both the H₆ pland H₆ fl coding cassettes were excised by NcoI/NheI digestion (an NheIsite existed at the 3' end of the RRS adaptor) and inserted intoNcoI/NheI digested pET11d. These two constructs, pET11d His₆ pl RRS andpET11d His₆ fl RRS, were transformed into competent BL21 DE3! bacteriafor expression of the recombinant proteins. BL21 DE3! contains arecombinant phage λ lysogen, DE 3, with a phage T7 RNA polymerase geneunder the transcriptional control of the lac UV5 promoter. T7 RNApolymerase gene expression is induced by the addition of IPTG(Isopropyl-B-D-thiogalactopyrano-side), which in turn leads to highlevel expression of the recombinant gene subcloned 3' of the pETvector's T7 gn 10 promoter.

The cDNAs encoding the group II allergens from D. pteronyssinus and D.farinae were also obtained from Dr. Wayne Thomas in plasmid form assubclones from λgt11 (Chua et al., Int Arch. Appl. Immun., 91:118-123(1990); Trudinger et al., Clin. Exp. Allergy, 21:33-37 (1991)). Theoriginal Der p II cDNA was subcloned from an M13RF plasmid into the pCAand pGEX vectors by Dr. Roland Buelow at ImmuLogic (Palo Alto). Dr.Thomas' group had supplied the Der f II cDNA as a subclone in pGEX. BothpGEX plasmids harboring group II cDNAs were used as templates for PCRamplification with the same 5' sense/3' antisense primer pair (FIG. 2b).The primers were designed to fuse an EcoRI site (GAATTC encoding theamino acids EF) in frame with the NH₂ terminus of the mature group IIproteins and place a PstI site 3' of the group II coding region. An MJResearch Thermal Controller with a program of 30×(94° C. 1 min./55°1min. 30 sec./72° C. 2 min.) was used in conjunction with reagents from aPerkin Elmer--Cetus Gene Amp kit for PCR amplification. The PCR productswere EcoRI/PstI digested and subcloned into EcoRI/PstI digestedM13mp19RF. DNA sequence analysis was performed to verify the sequence ofthe PCR products. Correct M13RF were EcoRI/PstI digested, their group IIcDNA inserts isolated, and subcloned into EcoRI/PstI digested pTrc99AHis₆ Amb a I.1 RRS (which served to exchange the group II cDNAs with theragweed cDNA (FIG. 33)).

The Der f II cDNA possessed a sequence polymorphism at position 54(i.e., threonine residue in place of isoleucine). To alter thispolymorphism, site directed mutagenesis of the T₅₄ residue in the Der fII cDNA was performed using a Muta-Gene kit from Bio-Rad Laboratories,based on the method of Kunkel, Proc. Natl. Acad. Sci. USA, 82:488-492(1985). The original M13mp19RF with the T₅₄ Der f II cDNA wastransformed into CJ236 bacteria, which tolerate the incorporation ofuracil in place of thymidine during DNA replication, and single strandedphage DNA prepared as template. A mutagenic 17 base pair primer (FIG.2b) was annealed to the phage template DNA. T4 DNA polymerase was usedto copy the template DNA primed by the mutagenic oligonucleotide, and T4DNA ligase served to seal gaps in the DNA strand. The reaction wastransformed into MV1190 bacteria, which are wild type for the editing ofuracil residues from DNA and therefore selectively replicate themutagenized (non-uracil containing) strand, and single stranded phageDNA prepared from recombinant (white) plaques. Several recombinants weresubjected to DNA sequence analysis and Der f II cDNAs with the correctedI₅₄ sequence subcloned as EcoRI/PstI fragments into EcoRI/PstI digestedpTrc99A His₆ Amb a I.1 RRS.

Since one previous work had shown that the pET11d vector was, in mostcases, capable of expressing recombinant proteins at higher levels thanthe pTrc99A vector, the two mite group II cDNAs were subcloned as H₆fusion proteins into T7 vector. pTrc ppA His₆ f II RRS and pTrc99A His₆p II RRS were Nco I/NheI digested, the cDNA inserts isolated, andsubcloned into NcoI/NheI digested pET11d His₆ p1 (FIG. 33). Recombinantplasmids were transformed into BL 21 DE3! bacteria for expression.

BL21 DE3 host bacteria harboring the pET11d mite allergen expressionconstructs were freshly streaked onto a BHI agar plate (3.7% wt./vol.Difco Brain Heart Infusion; 1.5% wt./vol. Difco agar) supplemented with200 μg/ml ampicillin and incubated overnight at 37° C. A single colonywas innoculated into a 2 ml of 200 μg/ml ampicillin/BH1 media (3.7%wt./vol. Difco Brain Heart Infusion) and shaken at 300 rpm at 37° C.until turbid but not saturated. The 2 ml culture was then added to 100ml of 200 μg/ml ampicillin/BH1 media, shaken at 300 rpm at 37° C. untilturbid but not saturated, at which point the culture was divided into18×500 ml (9 litres) of 200 μg/ml ampicillin/BH1 media and shaken at 300rpm at 37° C. When the OD₅₉₅ of the culture reached 1.0, expression ofthe recombinant molecules was induced by the addition of IPTG to 400 μM,and allowed to continue for two hours.

Bacteria were harvested by centrifugation at 10,000×g for 15 minutes,and resuspended in 1/20^(th) volume of lysis buffer (0.2 mg/ml lysozyme,100 mM NaPO₄ pH 8.0, 50 mM NaCl), incubated on ice for 30 minutes, andfrozen at -70° C. The frozen bacteria were fractured by a quick thaw at37° C. and then sonicated 5 times for 20 seconds at 30 second intervals.The sonicated samples were centrifuged at 15,000×g to separate solubleand particulate bacterial proteins. The soluble proteins were pouredoff, and the pelleted protein resuspended in 6M guanidine HCl, 100 mM2-mercaptoethanol, 100 mM NaPO₄, 10 mM Tris pH 8.0. This suspension wassubjected to centrifugation at 15,000×g, and the supernatant removed,adjusted to pH 8.0 with 10N NaOH, and applied to an NTA agarose columnthat had been equilibrated in 6M guanidine HCl, 100 mM NaPO₄, 10 mM TrispH 8.0. The column was washed in 6M guanidine HCl, 100 mM NaPO₄, 10 mMTris pH 8.0 until the OD₂₈₀ of the effluent reached background. Thecolumn buffer was then switched to 8M urea, 100 mM NaPO₄, 10 mM Tris pH8.0. After equilibration, a more stringent wash was performed in 8Murea, 100 mM NaOAc, 10 mM Tris pH 6.3 until the OD₂₈₀ of the effluentreached background. Recombinant mite protein (as an H₆ fusion) was theneluted in 8M urea, 100 mM NaOAc, 10 mM Tris pH 4.5 and collected inaliquots whose OD₂₈₀ profile was monitored. The protein peak wasdialyzed 3 times into 500 volumes of PBS for human T-cell analysis.Yield ranged from 10 to 70 mg of recombinant protein per liter withpurity (as determined by densitometric scanning) ranging from 80 (Der fII) to 95%.

To further purify the recombinant Der f II protein allergen, ReversePhase HPLC chromatography was applied. Approximately 100 mg of His₆ -Derf II protein was reduced in 20 mM DTT at 36° C. for 30 minutes and thenapplied to a Pharmacia PRO RPC HR 10/10 column in 0.1% vol./vol. TFA/H₂O. The flow rate for the column was 1.5 ml/min. with a gradient of 0-70%acetonitrile in 0.1% TFA over 40 minutes, followed by a gradient of70-100% acetonitrile in 0.1% TFA. His₆ -Der f II protein eluted between54-96% acetonitrile. Fractions detected at 214 nm and 280 nm wereanalyzed for purity by SDS-polyacrylamide gelelectrophoresis/densitometric scanning. Those fractions with His₆ -Der fII protein of purity greater than 95% were subsequently utilized forhuman T-cell mapping of the allergen. Yield from the preparative ReversePhase HPLC was approximately 69%.

Determination of Nucleotide Sequence Polymorphisms in the Der p I, Der pII and Der f II Allergens

It was expected that there were sequence polymorphisms in the nucleicacid sequence coding for Der p I, Der p II, Der f I and Der f₋₋ II, dueto natural allelic variation among individual mites. Several nucleotideand resulting amino acid sequence polymorphisms have been discoveredduring the sequencing of different Der p I,₋₋ Der p II and Der f IIclones. The amino acid sequence polymorphisms are shown in FIGS. 22, 23and 24.

EXAMPLE III Synthesis of Overlapping Peptides

Der p I, Der f I, Der p II, and Der f II overlapping peptides as shownin FIGS. 3 and 4 were synthesized using standard Fmoc/tBoc syntheticchemistry and purified by dialysis or Reverse Phase HPLC. The amino acidresidues of the synthesized peptides are in brackets by the peptide nameand the amino acid sequence (in single letter code) is next to thepeptide name. The peptide names are consistent throughout the Figures.The Der p I, Der f I, Der p II and Der f II proteins were divided intooverlapping peptides in such a way that the overlapping peptides of Derp I and Der f I as well as for Der p II and Der f II have correspondingamino acid residue numbers, e.g. DP I-1 and DF I-1 both contain aminoacid residues 1-20 of the Der p I and Der f I allergens, respectively.This correspondence in amino acid position between the Der p and Der fpeptides was done purposefully in order to best test forcross-reactivity of Der p and Der f T cell epitopes and, thus, determinepeptides which, upon administration to an individual sensitive to dustmite, would treat sensitivity to both Der p and Der f allergens. In thedesign of the overlapping peptides the relationship of the Group I andGroup II allergens at the level of T cell cross-reactivity wasconsidered. In addition, the function of the Group I allergens as serineproteases was considered and the amino acid sequences of other knownserine proteases, e.g., papain, actinidin, were examined to identifysimilar conserved and variable regions within the Group I allergens. Itwas expected that conserved regions within the Group I allergens wouldcontain "shared" T cell epitopes.

EXAMPLE IV Mite Allergic Patient Primary T Cell Responses to Der p I andDer p II Proteins and Peptides

Peripheral blood mononuclear cells (PBMC) were purified byFicoll-Hypaque centrifugation of a peripheral blood specimen frommite-allergic patient R.B. and were assayed for proliferation inresponse to various antigens, i.e., affinity-purified Der p I, affinitypurified Der p II, various Der p I and Der p II peptides. For assay,5×10⁴ PBMC were cultured in triplicate microwells for 7 days at 37° C.in the presence of various concentrations of antigen in 200 μl RPMI-1640containing 5% human AB serum. Each well then received 1 μCi tritiatedthymidine for 16 hours. The counts incorporated were collected ontoglass fiber filters and processed for liquid scintillation counting.Table I shows the results of this assay. The CPM+/- standard deviationare shown. The stimulation index of each response (S.I.) is the ratio ofthe ³ H-thymidine CPM incorporated by the cells in response to antigendivided by the ³ H-thymidine CPM incorporated by cells in medium only.The results indicate that this patient responds with an S.I. of at least2.0 to peptides DP I-1, DP I-3, DP I-8, DP I-10, DP I-5.2, DP II-4 andDP II-9. Thus, these peptides contain Der p I or Der p II T cellepitopes recognized by T cells from this particular allergic patient.

                  TABLE I                                                         ______________________________________                                                 Concentration                                                        Antigen  μg/ml      CPM ± S.D.                                                                           S.I.                                         ______________________________________                                        Medium   --            1071 ± 30                                                                            --                                           Der p I  10             920 ± 15                                                                            0.9                                                   30            2122 ± 93                                                                            2.0                                                   100           1492 ± 13                                                                            1.4                                          DP I-1   10            1099 ± 48                                                                            1.0                                                   30            3527 ± 73                                                                            3.3                                                   100           2746 ± 81                                                                            2.6                                          DP I-2   10            1395 ± 47                                                                            1.3                                                   30            1283 ± 34                                                                            1.5                                                   100           1486 ± 38                                                                            1.4                                          DP I-3   10            2608 ± 52                                                                            2.4                                                   30            1561 ± 13                                                                            1.5                                                   100           5252 ± 67                                                                            4.9                                          DP I-8   10            1439 ± 32                                                                            1.3                                                   30            1045 ± 32                                                                            1.0                                                   100           2272 ± 40                                                                            2.1                                          DP I-10  10            1936 ± 50                                                                            1.8                                                   30            3042 ± 89                                                                            2.8                                                   100           2644 ± 63                                                                            2.5                                          DP I-5.2 10            1374 ± 20                                                                            1.3                                                   30            2241 ± 87                                                                            2.1                                                   100           3132 ± 111                                                                           2.9                                          Der p II 10            1113 ± 35                                                                            1.0                                                   30            2104 ± 43                                                                            2.0                                                   100           1057 ± 27                                                                            1.0                                          DP II-4  10            2126 ± 25                                                                            2.0                                                   30            1979 ± 94                                                                            1.8                                                   100           2314 ± 116                                                                           2.2                                          DP II-9  10            3970 ± 87                                                                            3.7                                                   30            4464 ± 86                                                                            4.2                                                   100           2237 ± 53                                                                            2.1                                          ______________________________________                                    

EXAMPLE V T Cell Epitope Studies with Der p I

Peripheral blood mononuclear cells (PBMC) were purified byFicoll-Hypaque centrifugation of 60 ml of heparinized peripheral bloodfrom house dust mite-allergic individuals who exhibited clinicalsymptoms of mite allergy and who were skin test positive for house dustmite.

10⁷ PBMC from individual 543 were cultured in 10 ml RPMI-1640 containing5% pooled human AB serum and supplemented with glutamine, penicillin,streptomycin and HEPES buffer in the presence of 20 μg/ml purifiednative Der p I/ml at 37° C. for 7 days. Viable cells were then purifiedby Ficoll-Hypaque centrifugation and cultured for 2 additional weeks inRPMI-1640/5% AB serum containing 5 units recombinant human IL-2/ml and 5units recombinant human IL-4/ml. The resting T cells were then tested ina secondary proliferation assay to assess T cell responses to varioushouse dust mite proteins and peptides. For assay, 2×10⁴ resting T cellswere cultured in 200 μl of RPMI-1640/5% AB serum for 3 days at 37° C. inthe presence of 2×10⁴ autologous Epstein-Barr virus transformed B cells(20,000 Rads) as antigen presenting cells with various concentrations ofpurified native Der p I or synthetic Der p I peptides. Each well thenreceived 1 μCi tritiated thymidine for 16 hours. The counts incorporatedwere collected onto glass fiber filters and processed for liquidscintillation counting. Medium alone, acting as negative control,contained no allergen or peptide. The results of this experimentindicate that this particular patient responds with an S.I. of at least2.0 to several peptides derived from the Der p I protein, including DPI-1, DP I-2, DP I-4, DP I-11, DP I-5, DP I-13, DP I-15, DP I-6.1, DPI-8, DP I-9, DP I-16, DP I-10 and (data not shown).

The above procedure was followed with a number of other house dust miteallergic individuals except a) in individual cases, the length of timeof cultivation with IL-2 and IL-4 varied; b) in individual cases, the Tcells were primed with either purified native (N) or recombinant (R) Derp I protein at either 20 μg/ml or 10 μg/ml; and c) in individual cases,x-irradiated (3500 Rads) autologous PBMC were used as antigen presentingcells in the secondary proliferation assay. In addition, three peptides(DP I-11 (SEQ ID NO: 117), DP I-12 (SEQ ID NO: 118), and DP II-3 (SEQ IDNO: 119)) were found to contain a low number of conservative changesfrom the native sequence in their amino acid sequence. Three additionalpeptides (DP I-11.1 (SEQ ID NO: 13), DP I-12.1 (SEQ ID NO: 14) and DPII-3.1 (SEQ ID NO. 43) were synthesized with no changes from the nativesequence. Some T cell analysis was done with the original peptides(i.e., DP I-11, DP I-12 and DP II-3). Following T cell analysisconducted with the additional peptides (i.e., DP I-11.1, DP I-12.1 andDP II-3.1) no significant difference in mean S.I. or percentage ofpositive responses between the original peptides and the additionalpeptides was detected. Thus, the data from both groups of peptides waspooled.

Individual results were considered positive and used if the individualresponded to the Der p I protein and at least one peptide derived fromDer p I at an S.I. of 2.0 or greater. A summary of the results of 33positive experiments is shown in FIG. 37. The resting T cell linesprimed with recombinant or native Der p I-stimulated PBMC were testedfor reactivity to synthetic Der p I peptides. The maximum response foreach peptide in a titration of the antigen is expressed as the T cellstimulation index (S.I.). The S.I. is the counts-per-minute (CPM)incorporated by cells in response to the peptide divided by the CPMincorporated by cells in medium only. An S.I. value greater than thebackground level indicates that the peptide contains a T cell epitope.However, only individual S.I. values greater than or equal to 2.0 (aresponse two-fold or greater over background), referred to herein as"positive" results, were used in calculating mean T cell stimulationindices for each peptide for the patient or group of patients tested. Inparentheses above each bar on the histogram are the mean T cellstimulation indices calculated after discarding the highest and lowestpositive responses for each peptide to minimize the effect of extremeoutliers. The T cell stimulation index is calculated by:

    (CPM of T cell+APC+Antigen)/CPM of T cell+APC+Control

The bar represents the cumulative rank of the peptide response in thegroup of patients. To determine the cumulative rank, the 5 peptides withthe highest S.I. in each individual were determined and assigned anumerical rank in descending order, with 5 representing the strongestresponse. The ranks for each peptide were then summed in the group ofpatients to determine the cumulative rank for the peptide. Above eachbar is the percent of positive responses with an S.I. of at least 2 tothe peptide in the group of patients tested. Given the percent positiveand the mean T cell stimulation index, the positivity index (P.I.) foreach peptide can be calculated. The P.I. for each individual isdetermined by multiplying the mean S.I. by the percent of individuals,in a population of individuals sensitive to house dust mite (e.g.preferably at least 15 individuals, more preferably at least 30individuals or more) who responded with an S.I. of at least 2.0 to thatpeptide (e.g., for DP I-1 in FIG. 37, the P.I. would be about 343(73%×4.7). The P.I. therefore represents both the strength of a T cellresponse to a peptide (S.I.) and the frequency of a T cell response to apeptide in a population of individuals sensitive to house dust mite.FIG. 37 demonstrates that peptides DP I-1, DP I-2, DP I-3, DP I-4, DPI-5, DP I-6.1, DP I-7.1, DP I-9, DP I-16, and DP I-10 containsignificant regions of T cell reactivity in this panel of patients.

EXAMPLE VI T Cell Epitope Studies with Der f I

Experiments similar to those of Example V were performed to determinethe T cell-reactive areas of the Der f I protein. For example, PBMC fromhouse dust mite-allergic patient 783 were isolated as described inExample V and were stimulated in vitro₋₋ with recombinant purified Der fI at 20 μg/ml. The results of the proliferation assay with Der f Ipeptides using x-irradiated (3500 Rads) autologous PBMC as antigenpresenting cells indicate that T cells from this patient respond to thepeptides DF I-8.1, DF I-9, DF I-6, DF I-10, DF I-2.1, DF I-3, DF I-11,DF I-5, DF I-1, and DF I-17 (data not shown).

The above procedure was followed in a number of patients except inindividual cases, T cell lines were primed with affinity purified Der fI at 20 μg/ml or at 10 μg/ml and x-irradiated (25,000 Rads) autologousEpstein-Barr virus transformed B cells were used as antigen presentingcells. A summary of the results of 16 positive experiments is shown inFIG. 6. The data was analyzed as described in Example V. The dataindicate that significant areas of T cell reactivity in the Der f Iprotein are found in the peptides DF I-1, DF I-2, DF I-3, DF I-4, DFI-11, DF I-5, DF I-6, DF I-7, DF I-14, DF I-15, DF I-8.1 and DF I-9.

EXAMPLE VII A Study Indicating the Cross-reactivity of Der p I and Der fI T Cell Epitopes

The Der p I and Der f I proteins are very homologous (81% identity).Thus, experiments, similar to those of Example V, were carried out todetermine the T cell responses of Der p I primed T cell lines whenchallenged with various Der p I peptides and substantially matching Derf II peptides. T cell lines were primed in vitro as described in ExampleV and tested for response to a set of substantially matching Der p I andDer f I peptides (e.g., DP I-1 (amino acid residues 1-20 of Der p I) orDF I-1 (amino acid residues 1-20 of Der f I). The data was analyzed asdescribed in Example V except the highest and lowest S.I. values of thepositive responses to each peptide were not omitted from the mean S.I.calculations. A summary of a number of such experiments is shown in FIG.7. The results of 14 positive experiments indicate that Der p I-primed Tcells respond to various Der f I peptides indicating cross-reactivitywithin the Group I allergens. Der p I primed T cells respondsignificantly to peptides DF I-1, DF I-2, DF I-3, DF I-4, DF I-11, DFI-12, DF I-15, DF I-8, DF I-9, DF I-6. In some patients, the Der f Ipeptide was a more potent stimulator of Der p I-primed T cells than thecorresponding Der p I peptide. FIG. 8 shows the results of inverseexperiments in which T cells from a number of patients were primed invitro to the Der f I protein and analyzed for response to various Der pI peptides and a set of substantially matching Der f I peptides. Theresults of 8 positive experiments indicate that Der f I primed T cellsrespond significantly to peptides DP I-1, DP I-3, DPI-4, DP I-11/11.1,DP I-14, DPI-5, DPI-15, and DP I-8.

EXAMPLE VIII T Cell Epitope Studies with Der p II.

Experiments similar to those of Example V were performed to determinethe T cell-reactive areas of the Der p II protein. For example, PBMCfrom house dust mite-allergic patient 348 were isolated as described inExample V and were stimulated in vitro with 20 μg/ml purified native Derp II. The results of a proliferation assay using x-irradiated (25,000Rads) Epstein-Barr virus transformed autologous B cells asantigen-presenting cells with various Der p II peptides demonstrate thatthis particular patient responds well to peptides DP II-1, DP II-7, DPII-8, DP II-2, and DP II-9 (data not shown).

The above procedure was followed with a number of patients except inindividual cases, T cell lines were primed with recombinant Der p II at20 μg/ml or at 3 μg/ml and x-irradiated (3500 Rads) autologous PBMC wereused as antigen-presenting cells. A summary of the results of 26positive experiments is shown in FIG. 9. The data was analyzed asdescribed in Example V, except the ranked sum of peptide responses wasanalyzed assigning a value of 3, 2 or 1 to the three highest S.I.responses. Areas of significant T cell reactivity within the Der p IIprotein for this panel of patients are found in peptides DP II-1, DPII-2, DP II-3, DP II-4, DP II-7, DP II-8 and DP II-9.

EXAMPLE IX T Cell Epitope Studies with Der f II

Experiments similar to those of Example V were performed to determinethe T cell-reactive areas of the Der f II protein. For example, PBMCfrom house dust mite-allergic patient 384 were stimulated in vitro withpurified recombinant Der f II and the T cell line was then challenged inthe presence of x-irradiated (25,000 Rads) autologous Epstein-Barr virustransformed B cells as antigen presenting cells with various overlappingDer f II peptides. The results of this proliferation assay indicate thatT cells from this particular patient respond well to peptides DF II-1,DF II-2, DF II-3.1, DF II-4.5, DF II-15, DF II-16, and DF II-19.1 (datanot shown).

The above procedure was followed with 10 patients, except in individualcases, T cell lines were primed by stimulating the patient PBMC with 20μg/ml or 3 μg/ml purified native Der f II, and were assayed in thepresence of x-irradiated (3500 Rads) autologous PBMC asantigen-presenting cells. A summary of the results of 10 positiveexperiments is shown in FIG. 10. The data was analyzed as detailed inExample IX, except the highest and lowest S.I. values of the positiveresponses to each peptide were not omitted from the calculations. Thedata indicate that significant areas of T cell reactivity within the Derf II protein are found in peptides DF II-1, DF II-2, DF II-13.1, DFII-4.5, DF II-15, DF II-17, and DF II-19.1.

EXAMPLE X A Study Indicating the Cross-Reactivity of Der p II and DerfII T Cell Epitopes

A study similar to that described in Example VII was carried out todetermine the T cell cross-reactivity of the Der p II and Der f IIproteins. T cells primed with the Der f II protein were challenged withvarious Der f II peptides and a set of substantially matching Der p IIpeptides. A summary of the results of 10 positive experiments is shownin FIG. 11. The results indicate that Der f II primed T cells respondsignificantly to peptides DP II-1, DP II-3/3.1, DP II-4, and DP II-7.FIG. 12 shows the results of inverse experiments in which T cells from anumber of patients were primed in vitro to the Der p II protein andanalyzed for response to various Der f II peptides. The results of 26positive experiments indicate that Der p II primed T cells respondsignificantly to peptides DF II-1, DF II-4.5, DF II-15, DF II-17, DFII-18 and DF II-19.1.

EXAMPLE XI Synthesis of Dominant Peptides

Based on the analyses described in Examples V-X, major areas of T cellreactivity within Der p I, Der p II, Der f I and Der f II wereidentified. In each study, all of the patients tested responded to theprotein allergen (e.g., Der p I) and at least one peptide derived from amajor area of T cell reactivity within the protein. Seven regions(Region 1, Region 2, Region 3, Region 4, Region 5, Region 6a and Region6b) of major T cell reactivity were identified in the Der p I and Der fI proteins. These regions are defined as follows: Region 1, amino acidresidues 1-28 of the Der p I and Der f I proteins; Region 2, amino acidresidues 36-68 of the Der p I and Der f I proteins; Region 3, amino acidresidues 74-109 of the Der p I and Der f I proteins; Region 4, aminoacid residues 118-139 of the Der p I and Der f I proteins; Region 5,amino acid residues 141-166 of the Der p I and Der f I proteins; andRegion 6a, amino acid residues 161-185 of the Der p I and Der f Iproteins and Region 6b, amino acid residues 173-201 of the Der p I andDer f I proteins.

Similarly, four regions of major T cell reactivity (Region 7, Region 8,Region 9, and Region 10) were identified in the Der p II and Der f IIproteins. These regions are defined as follows: Region 7, amino acidresidues 1-26 of the Der p II and Der f II proteins; Region 8, aminoacid residues 33-67 of the Der p II and Der f II proteins; Region 9,amino acid residues 79-104 of the Der p II and Der f II proteins; andRegion 10, amino acid residues 107-129 of the Der p II and Der f IIproteins. Based in part on the T cell reactivity described in ExamplesV-X, peptides derived from Der p I, Der f I, Der p II, and Der f II wereselected and modified by addition or deletion of amino acid residues ateither the 5' or 3' end of the peptide. In designing these selectedpeptides, various factors were considered, including the ranked sum ofthe overlapping peptides, the percentage of responses with an S.I. of atleast 2.0 to the peptides, the potential cross-reactivity of thepeptides, the difficulty of manufacture of the peptides, etc. T cellstudies similar to those described in Examples V-X were performed usingthese selected peptides to more precisely define the major areas of Tcell reactivity within Regions 1-6a and 6b of the Der p I and Der f Iprotein and Regions 7-10 of the Der p II and Der f II protein.

The results of T cell studies using selected peptides derived from theDer p I, the Der f I, the Der p II and the Der f II proteins are shownin FIGS. 13-18a-d. The procedure described in Example V was followedwith T cell lines from a number of patients primed in vitro to the Der pI protein then analyzed for response to selected peptides derived fromthe Der p I sequence. The results of 33 positive experiments shown inFIG. 13 indicate that the Der p I primed T cells respond significantlyto peptides found in peptides DP I-21.1, DP I-21.2, DP I-22.2, DPI-25.2, DP I-22.1, DP I-23.1, DP I-23.2, DP I-26.1, and DP I-28.1.

Similarly, the procedure described in Example VI was followed with Tcell lines from 9 patients primed in vitro to the Der f I protein thenanalyzed for response to selected peptides derived from the protein. Thedata was analyzed as described in Example VI. The results of the 9patients shown in FIG. 14 demonstrate T cell reactivity to selectedpeptides from Der f I.

In another experiment, the T cell lines from a number of patients wereprimed in vitro to the Der p I protein and analyzed for response toselected peptides derived from the Der p I protein and a set ofsubstantially matching peptides derived from the Der f I protein. Thedata from 30 positive experiments was analyzed as described in ExampleV. As shown in FIG. 15a, the Der p I primed T cells respondsignificantly to peptides DF I-21.1, DF I-21.2, DF I-23.1, DF I-22.2, DFI-22.3, DF I-22.4, DF I-23.2, DF I-25.1, DF I-26.1 and DF I-27.1. FIG.15b is a subset of the data shown in FIG. 15a and shows the response ofDer p I primed T cells to peptides analyzed by ranked sum. FIG. 15bshows that DPI-23.1 has the highest ranked sum of this group of peptidesin this study. FIG. 16a shows the results of the inverse experiment inwhich T cells from 9 patients were primed in vitro to the Der f Iprotein and challenged with selected Der f I peptides and a set ofsubstantially matching Der p I peptides. The results indicate that Der fI primed T cells from 9 patients respond to selected peptides from Der pI. FIG. 16a shows that DF I-22.1 and DF I-25.1 have the higheststimulation indexes of this group of peptides. FIG. 16b is a subset ofthe same data as shown in FIG. 16a and shows the response of preferredDer f I and Der p I peptides is shown. FIG. 16b shows that DFI-22.2 hasthe highest stimulation index of this group of preferred peptides inthis experiment.

Another experiment following the procedure described in Example VIIIanalyzed the response of 29 patients primed in vitro to the Der p IIprotein and challenged with selected peptides derived from the Der p IIsequence. FIG. 17a shows that Der p II primed T cells from one patientrespond to selected peptides from Der p II. FIG. 17b shows the resultsfrom an experiment similar to that shown in FIG. 17a with a set of 30patients and with the high and low omitted from the mean. FIG. 17b showsthat DPII-20.6 has the highest ranked sum of this group of preferredpeptides in this study. FIG. 18a shows the inverse experiment in which Tcells from 1 patient were primed in vitro to the Der f II protein andchallenged with selected peptides derived from the Der p II sequence.FIG. 18a shows that Der f II primed T cells from 10 patients respond toselected peptides from Der p II. As shown in FIG. 18a DP II-25 has thehighest stimulation index. FIG. 18b is a subset of the same data shownin FIG. 18a and shows the response of native Der f II primed T celllines to preferred Der p II peptides analyzed by ranked sum. As is shownin FIG. 18b, DPII-25.2 has the highest ranked sum of the preferredpeptides in this study.

EXAMPLE XII Study Indicating Cross-reactivity of Selected Group I andGroup II Epitopes

A study similar to that described in Example VII was carried out with Tcells from 4 matched patients primed in vitro with Group I proteins fromDer f and Der p then analyzed for response to selected preferredpeptides from Der p I and Der f I. The results in FIG. 18c demonstratethat T cell reactivity to a number of the selected preferred Der p Ipeptides was essentially equivalent for their Der f I counterpart andvice-versa.

FIG. 18d shows the results of a similar study with T cells from 6matched patients primed in vitro with Group II proteins from Der p andDer f, then analyzed for response to selected preferred Der p II and Derf II peptides. Similar to the results in FIG. 18c, T cell reactivity toa number of the selected preferred peptides of₋₋ Der p II areessentially equivalent to their Der f II counterparts and vice-versa.

EXAMPLE XIII Direct Binding Assay of IgE to Mite Allergen Proteins andPeptides

Corning assay plates (#25882-96) were coated with 5 μg/ml of eachcoating antigen listed in FIGS. 19, 20 and 21 at 50 μl/100 ml/well andincubated overnight at 4° C. The coating antigens were removed and thewells are blocked with 0.5% gelatin in PBS, 300 μl/well for 2 hours atroom temperature. Pooled human plasma (a mix of plasma samples from 20patients that were skin test positive for commerical mite extract) wasserially diluted with PBS-Tween 20 (PBS with 0.05% nonionic detergentTween-20 Sigma, St. Louis, Mo.) and 100 μl/well was added and incubatedovernight at 4° C. (plasma dilutions were tested in duplicate). Thesecond antibody (biotinylated goat anti-Human IgE, 1:1000, Kirkegaard &Perry Laboratories Inc, Gaithersburg, Md.), was added at 100 μl/well forone hour at room temperature. This solution was removed andstreptavidin-HRPO, 1:10000, (Southern Biotechnology Associates, Inc.,Birmingham, Ala.) was then added at 100 μl/well for one hour at roomtemperature (all wells are washed three times with PBS-Tween betweeneach incubation step). TMB Membrane Peroxidase Substrate system(Kirkegaard & Perry Laboratories) was freshly miced, and added at 100ml/well. The color was allowed to develop for 2-5 minutes. The reactionwas stopped by the addition of 100 ml/well of 1M phosphoric acid. Plateswere read on a Microplate IL310 Autoreader (Biotech Instruments,Winooski, Vt.) with a 450 nm filter. The absorbance levels of duplicatewells were averaged. The graphed results (log of the dilution versusabsorbance) of the ELISA assays are shown in FIGS. 19a-b, 20a-b and21a-h. The order of coating antigens listed vertically in these figurelegends corresponds in orDer from left to right to the coating antigenslisted for each histogram.

The results of the ELISA assay shown in FIG. 19b demonstrate goodbinding of both biochemically purified Der p II and recombinant Der p II(rDer p II) with human IgE and no detectable binding to the Der p IIpeptides. The IgE binding to the Der p set of peptides and proteins(FIGS. 20a and 20b) shows the same pattern of reactivity as the Der fset. That is, no detectable binding to Der f I or Der f II peptides orrecombinant Der f I with binding to only biochemically purified Der fIand recombinant and biochemically purified Der f II. In both cases thereappears to be better binding to recombinant Der p II or Der f₋₋ II thanto the biochemically purified forms. All the conclusions derived fromthe above ELISA assay data were corraborated by another assay method,dot blots on nitrocellulose paper, using the same set of antibody andantigen reagents.

The antigen preparation that was used as a positive control was amixture of the four major biochemically purified mite allergens (termPMA for Purified Mite Allergen); Der f I, Der f II, Der p I and Der pII. The stock was generated at a concentration of 100 μgs of eachprotein per millimeter or 400 μgs total protein/ml. This preparation wasused on each coated ELISA plate. The results from these ELISA assays areshown in FIG. 21a-h. There is clear binding to either the purified orrecombinant protein or the PMA antigen preparation on each plateindicating good IgE reactivity. However, the PMA antigen preparation,does exhibit a high degree of non-specific reactivity shown in thebackground dilution where no first antibody solution was added. Thisnon-specific reactivity occurs between the PMA antigen and thebiotinyulated second antibody and does not compromise the finding ofspecific IgE reactivity to the antigen. Using a quantitative value oftwo-fold over background at the highest plasma concentration as apositive reading, there is no detectable IgE reactivity to any one ofthe 56 peptides screened by this assay method.

EXAMPLE XIV T Cell Epitope Studies with Modified Peptides Synthesis ofPeptides

Modified Der p I, Der p II, and Der f II peptides were synthesized usingstandard Fmoc/tBoc synthetic chemistry and purified by Reverse PhaseHPLC. FIG. 30 shows the modified house dust mite peptides used in thesestudies. These modified peptides were constructed for enhancedsolubility as compared to the peptides from which they were derived. Inalmost all cases, these modified peptides exhibited enhanced solubilityas compared to the peptides from which they were derived (data notshown). Other peptides used in these studies and which are shown inFIGS. 31a-b and 32a-b may be found in FIGS. 3 and 4 and are discussed inExamples III-IX and XI. The peptide names are consistent throughout.

T Cell Responses to Cedar Pollen Antigen Peptides

Peripheral blood mononuclear cells (PBMC) were purified by lymphocyteseparation medium (LSM) centrifugation of 60 ml of heparinized bloodfrom house dust mite allergic patients who exhibited clinical symptomsof seasonal rhinitis and were MAST and/or skin test positive for housedust mite protein allergen. Long term T cell lines were established bystimulation of 2×10⁶ PBL/ml in bulk cultures of complete medium(RPMI-1640, 2 mM L-glutamine, 100 U/ml penicillin/streptomycin, 5×10⁻⁵ M2-mercaptoethanol, and 10 mM HEPES supplemented with 5% heat inactivatedhuman AB serum) with 30 μg/ml of purified native Der p I or Der p II for5-6 days at 37° C. in a humidified 5% CO₂ incubator to select for housedust mite protein allergen reactive T cells. This amount of primingantigen was determined to be optimal for the activation of T cells fromhouse dust mite allergic patients. Viable cells were purified by LSMcentrifugation and cultured in complete medium supplemented with 5 unitsrecombinant human IL-2/ml and 5 units recombinant human IL-4/ml for upto three weeks until the cells no longer responded to lymphokines andwere considered "rested". The ability of the T cells to proliferate toselected peptides, purified Der p I or Der p II, or positive (PHA)controls or negative controls (medium only) was then assessed. Forassay, 2×10⁴ rested cells were restimulated in the presence of 2×10⁴autologous Epstein-Barr virus (EBV)-transformed B cells (prepared asdescribed below) (gamma-irradiated with 25,000 RADS) or with 5×10⁴autologous PBMs (gamma-irradiated with 3500 RADS), with 2-50 μg/ml ofpurified native Der p I or Der p II in a volume of 200 μl completemedium in duplicate or triplicate wells in 96-well round bottom platesfor 3 days. Each well then received 1 μCi tritiated thymidine for 16-20hours. The counts incorporated were collected onto glass fiber filtermats and processed for liquid scintillation counting. Titrations using Tcells from one individual were conducted which showed the effect ofvarying antigen dose in assays with purified native Der p I or Der p IIand several of the peptides synthesized as described above. Thetitrations were used to optimize the dose of peptides in T cell assays.

The maximum response in a titration of each peptide is expressed as thestimulation index (S.I.). The S.I. is the counts per minute (CPM)incorporated by cells in response to peptide, divided by the CPMincorporated by cells in medium only. An S.I. value equal to or greaterthan 2 times the background level is considered "positive" and indicatesthat the peptide contains a T cell epitope. The positive results wereused in calculating mean stimulation indices for each peptide for theindividual patient tested.

The above procedure was followed with 18 patients in the studies usingpeptides derived from Der p I and Der f I (FIG. 31a-b), and 21 patientsin the studies with Der p II peptides (FIGS. 32a-b). Individual patientresults were used in calculating the mean S.I. for each peptide if thepatient responded to the Der p I or Der p II protein at an S.I of 2.0 orgreater and the patient responded to at least one peptide derived from ahouse dust mite allergen at an S.I. of 2.0 or greater. A summary ofpositive experiments from the patients tested is shown in FIGS. 32a and32b and FIGS. 33a and 33b. The bars represent the positivity index.Above each bar is the percent of positive responses with an S.I. of atleast 2.0 to the peptide or protein in the group of patients tested. Inparenthesis above each bar are the mean stimulation indices for eachpeptide or protein for the group of patients tested.

As shown in FIG. 31a, all eighteen T cell lines responded positively tomodified peptides DP I-23.31 (SEQ ID NO:165), DP I-23.35 (SEQ IDNO:167), and DP I-26.6 (SEQ ID NO:168) when primed with affinitypurified Der p I, and as shown in FIG. 32a, all 21 T cell linesresponded positively to modified peptides DP II-20.9 (SEQ ID NO:169), DPII-20.11 (SEQ ID NO:169), DP II-20.10 (SEQ ID NO:170), DP II-20.8 (SEQID NO:171), DP II-22.19 (SEQ ID NO:172), DP II-22.22 (SEQ ID NO: 175),DP II-22.23 (SEQ ID NO:177), DP II-22.14 (SEQ ID NO:180), and DP II-25.9(SEQ ID NO: 183), when primed with Der p II indicating that all of themodified peptides tested contain at least one T cell epitope. The otherpeptides in FIGS. 31a-b and 32a-b were previously tested (see, ExamplesIII-IX and XI) and were used in these studies as controls.

Preparation of (EBV)--transformed B Cells for Use as Antigen PresentingCells

Autologous EBV-transformed cell lines were γ-irradiated with 25,000 Radand used as antigen presenting cells in secondary proliferation assaysand secondary bulk stimulations. These EBV-transformed cell lines weremade by incubating 5×10⁶ PBL with 1 ml of B-59/8 Marmoset cell line(ATCC CRL1612, American Type Culture Collection, Rockville, Md.)conditioned medium in the presence of 1 μg/ml phorbol 12-myristate13-acetate (PMA) at 37° C. for 60 minutes in 12×75 mm polypropyleneround-bottom Falcon snap cap tubes (Becton Dickinson Labware, LincolnPark, N.J.). These cells were then diluted to 1.25×10⁶ cells/ml inRPMI-1640 as described above except supplemented with 10%heat-inactivated fetal bovine serum and cultured in 200 μl aliquots inflat bottom culture plates until visible colonies were detected. Theywere then transferred to larger wells until the cell lines wereestablished.

EXAMPLE XV Direct binding assay if IgE to Modified Der p and Der fPeptides

Plasma samples from 19 mite-allergic patients (4+skin test) and a poolof plasma from 18 additional patients were tested using the procedureoutlined in Example XIII with minor differences (Costar ELISA plateswere used instead of Corning, and 10 μl/ml peptides were coated onto theplates instead of 5 μl/ml. In each case, a control was run for IgEbinding to uncoated plates (no peptide). No plasma samples contained IgEto any of the peptides. All samples did contain IgE to Der p I and/orDer p II. The modified peptides tested were, DP II-20.8 (SEQ ID NO:171), DP II-20.9 (SEQ ID NO: 169), DPII-20.10 (SEQ ID NO: 170), DPII-20.11 (SEQ ID NO: 169), DP I 23.31 (SEQ ID NO:165), DP I 23.35 (SEQID NO:167), DP II-22.14 (SEQ ID NO: 180), DP II-22.19 (SEQ ID NO:172),DP II-22.22 (SEQ ID NO:175), DP II-22.23 (SEQ ID NO:177), and DP II-25.9(SEQ ID NO:183)

EXAMPLE XVI Analysis of IgE -Producing B Lymphocytes in Atopic andNon-Atopic Individuals

The factors influencing the progression of a normal immune response toforeign antigen involving IgM to IgG to an allergic response involvingIgE, are unknown, as are the properties of allergens that induce thistransition. Although Burastero et al., J. Allergy Clin. Immunol.,91:1075-1081 (1993) reported that the atopic subjects had significantlyhigher frequencies of Der p-specific T cells than did healthy subjects.both O'Heir et al., Immunology, 66:499-504 (1989) and Wierenga et al.,Eur. J. Immunol., 20:1519-1526 (1990) have cloned T cells specific forDer p I and Der p II allergens from both atopic and non-atopic donors,supporting the current findings that both atopic and non-atopic subjectsmount an antibody response to Der p. Only those clones isolated frommite-allergic subjects however, could be induced by mite allergens toprovide help for IgE synthesis by autologous B lymphocytes (O'Heir etal, (supra I and Wierenga et al., (supra)) Furthermore, Parronchi etal., Proc. Natl. Acad. Sci. USA, 88:4538-4542 (1991) found that onlyallergen specific clones from atopic subjects produced IL4 and IL5 andprovided help for IgE synthesis, whereas tetanus toxoid-specific dinesisolated from the same subject produced IL4 and IFN-γ, and were unableto provide help for IgE synthesis. Taken together, these studies suggestthat T lymphocytes of atopic subjects respond to allergens differentlythan those of non-atopic subjects, producing a panel of lymphokines(high IL4, low IFN-γ) that favors the production of IgE (Pene et al.,Proc. Natl. Acad. Sci. USA, 85:6880-6884 (1988). Because IL4 enhancesIgE production by directing isotype switch recombination to the Cε locus(Gascan et al. J. Exp. Med., 173:747-750 (1991), such a mechanism wouldresult in an increased frequency of IgE-producing B cell precursors, andmay help to account for the differences in precursor frequency observedin the studies described below. The specific genetic, regulatory, orenvironmental differences that underlie the distinct T cell responses toallergen in atopic and non-atopic subjects remain to be elucidated.

The frequency of B cell precursors in atopic and non-atopic subjectscommitted to the production of IgE, IgM and IgG was directly compared bylimiting dilution analysis of EBV transformants. Although no differencewas found in the IgM or IgG-producing B cell precursor frequency, theatopic subjects had a significantly higher frequency of B lymphocytescommitted to the production of IgE (0.79% of total Ig-producing cells,as compared to 0.15% for the control group; p<0.005). This differencewas associated with significantly greater total plasma IgE concentrationin the atopic group (1.08 μg/ml) as compared to non-atopic controlsubjects (0.110 μg/ml)(p<0.005), suggesting that one factor contributingto the high plasma IgE titers in atopic subjects may be the increasedfrequency of IgE-producing B lymphocytes. The titers ofallergen-specific IgE, IgM, and IgG in the plasma of atopic andnon-atopic subjects were also compared. Plasma samples from 12 atopicand 12 non-atopic subjects were assayed for levels of total IgM, IgG,and IgE using specific ELISAs (direct ELISA, competitive inhibitionELISA). Although only the atopic subjects produced IgE directed againstthe allergy to which they were sensitive, atopic and non-atopic subjectshad comparable plasma titers of IgM and IgG specific for the major housedust-mite allergens, Der p I and Der p II, or the major ragweedallergen, Amb a I. This finding was supported by the similar frequenciesof allergen-specific IgM-producing B lymphocytes in the atopic andnon-atopic subjects. Thus, although all exposed individuals undergo animmune response to the allergen involving the production of specific IgMand IgG, only in atopic subjects is the response extended to includespecific IgE. The evidence thus indicates that the higher frequency ofIgE-producing B lymphocytes and the higher titers of total IgEcharacteristic of allergic subjects may be indicative of a regulatoryenvironment that favors the production of allergen-specific IgE.

EXAMPLE XVII Determination of the Percentage of Total House Dust MiteGroup I T Cell Reactivity and Patient Coverage of the Combination ofUnique Der p I and Der f I Peptides.

Synthesis of Overlapping Peptides

Der p I was divided into a set of 17 overlapping peptides. Overlappingpeptides and Der p I and Der f I unique peptides (i.e. DPI-21.2,DFI-23.31, DPI-26.6) were synthesized using standard Fmoc/tBoc syntheticchemistry and purified by Reverse Phase HPLC. FIG. 33 shows the uniqueDer p I, and Der f I peptides and FIG. 34 shows the overlapping Der p Ipeptides used in these studies. The peptide names are consistentthrougout.

T cell responses to Group I overlapping peptides and unique peptides

Peripheral blood mononuclear cells (PBMC) were purified byFicoll-Hypaque centrifugation of 60 ml of heparinized peripheral bloodfrom house dust mite-allergic individuals who exhibited clinicalsymptoms of mite allergy and who were skin test positive for house dustmite.

10⁷ PBMC fro each patient were cultured in 5 ml RPMI-1640 containing 5%pooled human AB serum and supplemented with glutamine, penicillin,streptomycin and HEPES buffer in the presence of 20 μg/ml purifiednative Der p I/ml at 37° C. for 6 days. Viable cells were then purifiedby Ficoll-Hypaque entriguation and cultured for 2-3 additional weeks inRPMI-1640/5% AB serum containing 5 units recombinant human IL-2/ml and 5units recombinant human L-4/ml. The resting T cells were then tested ina secondary proliferation assay to assess T cell responses to purifiednative Der p I, overlapping peptides and unique peptides. For assay,2×10⁴ resting T cells were cultured in 200 μl of RPMI-1640/5% AB serumfor 3 days at 37° C. in the presence of 2×10⁴ autologous Epstein-Barrvirus transformed B cells (20,000 Rads) or in the presence of 5×10⁴ PBMC(3500 Rads) as antigen presenting cells with various concentrations ofpurified native Der p I or synthetic Der p I unique or peptide oroverlapping peptides. Each well then received 1 μCi tritiated thymidinefor 16 hours. The counts incorporated were collected onto glass fiberfilters and processed for liquid scintillation counting. Medium alone,acting as negative control, contained no allergen or peptide. Theresults are shown in FIG. 35. The highest stimulation index greater thanor equal to 2.0 in response to each peptide was recorded for eachsubject tested. The data were then analyzed by the equations describedearlier in the specification.

The combination of Group I candidate peptides DPI-21.2, DFI-22.2,DPI-23.31, DPI-26.6 had a range of T cell reactivity of about 38-67 &based on an analysis of 39 patients, the frequency of response at 82%represents reactivity to at least one of the candidate peptides,indicating that this combination of peptides fits the first criteria for"unique" peptides of the invention in that the combination of peptidesDPI-21.2, DFI-22.2, DPI-23.31, DPI-26.6 comprise a sufficient percentageof the total T cell reactivity to Group I protein allergens from Der pand Der f in a substantial percentage of the population tested.

EXAMPLE XVIII Determination of the Percentage of Total House Dust MiteGroup II T Cell Reactivity and Patient Coverage of the Combination ofUnique Der p II Candidate Peptides

Synthesis of Overlapping Peptides

Der p II was divided into a set of 9 overlapping peptides. OverlappingDer p II peptides and Der p II unique peptides (i.e. DPII-20.9 (SEQ. ID.NO. 169), DPII-22.14 (SEQ. ID. NO. 180), and DPII-25.15 (SEQ. ID. NO.188)) were synthesized using standard Fmoc/tBoc synthetic chemistry andpurified by Reverse Phase HPLC. FIG. 33 shows the unique Der p IIpeptides and FIG. 34 shows the overlapping Der p II peptides used inthese studies. The peptide names are consistent throughout.

T cell responses to Group II overlapping peptides and unique candidatepeptides

Secondary Der p II reactive T cell cultures derived from 30mite-allergic patients were analyzed for reactivity to an overlappingset of Der p II peptides and Der p II candidate peptides. in an in vitroT cell proliferation assay as described in Example XVII. The results areshown in FIG. 36. The highest stimulation index greater than or equal to2.0 in response to each peptide was recorded for each subject tested.The data were then analyzed by the equations described earlier in thespecification.

The combination of candidate peptides DPII-20.9 (SEQ. ID. NO. 169),DPII-22.14 (SEQ. ID. NO. 180), and DPII-25.15 (SEQ. ID. NO. 188), had arange of T cell reactivity of about 36%-51% based on an analysis of 30patients (FIG. 36). The frequency of response to at least one of thecandidate peptides was about 63%, indicating that this combination ofpeptides fits the first criteria for "unique" peptides of the invention,in that the combination of Der p II peptides DPII-20.9 (SEQ. ID. NO.169), DPII-22.14 (SEQ. ID. NO. 180), and DPII-25.15 (SEQ. ID. NO. 188),comprise a sufficient percentage of the total T cell reactivity to Der pII in a substantial percentage of the population tested.

EXAMPLE XVIX Determination of pH-solubility and pH-stability Profiles ofCandidate Peptides of the Invention

1. Buffer Preparation

50 mM sodium phosphate stock solutions:

Stock solution A: 0.66 g (0.05 mol) of monobasic sodium phosphatemonohydrate U.S.P. and 50 mg EDTA disodium dihydrate, U.S.P. weredissolved in 100 mL of WFI. The solution was filtered through a 0.2micron filter

Stock solution B: 0.71 g (0.05 mol) of dibasic sodium phosphate U.S.P.were disolved in 100 ml WFI. The solution was filtered through a 0.2micron filter.

2. Initial peptide dispersions

Dispersion A: 3.0 mg of each candidate peptide,DPI-21.2 (SEQ. ID. NO.28), DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165), DFI-26.6(SEQ. ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14 (SEQ. ID.NO. 180) and DPII-25.15 (SEQ. ID. NO. 188), was weighed out separatelyand placed in separate 1.5 mL eppendoff vials with 600 μL of stocksolution A. The composition was agitated for 5 seconds to mix well.

Dispersion B: 3.0 mg of each peptide, DPI-21.2 (SEQ. ID. NO. 28),DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165), DFI-26.6 (SEQ.ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14 (SEQ. ID. NO.180) and DPII-25.15 (SEQ. ID. NO. 188), was weighed out separately andplaced in separate 1.5 mL eppendoff vials with 600 μL of stock solutionB. The mixture was agitated for 5 seconds to mix well.

Dispersions A and B were sonicated for 2 minutes for good homogeneity. Asmall volume was pipetted from each dispersion into a labeled eppendoffvial according to the following volume ratio:

    ______________________________________                                        Vial Suspension A                                                                             Suspension B                                                                             Total Volume                                                                           Estimated final                           #    (μL)    (μL)    (μL)  pH                                        ______________________________________                                        1    100         0         100      5.2                                       2    80         20         100      6.2                                       3    60         40         100      6.6                                       4    40         60         100      6.8                                       5    20         80         100      7.1                                       6     0         100        100      8.0                                       ______________________________________                                    

The resultant solutions/suspensions were stored in the dark at about 22°C.±2 for 24 hours without agitation. The solutions were filtered andfiltrates were analyzed for pH and peptide concentration.

The concentration of filtered peptide solutions was determined by HPLCanalysis. In this experiment, the solubility of peptide at each pH isdefined as the amount of peptide remaining in solution after filtrationthrough a membrane filter having a 0.2 micrometer pore size. The extentof degradation of peptides was estimated by calculating the percent oftotal degradant peak area over the total peak area.

The pH values with respect to the solubility values was plotted and areshown in FIG. 37 for each respective peptide. As shown in the solubilitycurve for each peptide as represented in FIG. 37, peptides DPI-21.2(SEQ. ID. NO. 28), DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO.165), DFI-26.6 (SEQ. ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169),DPII-22.14 (SEQ. ID. NO. 180) and DPII-25.15 (SEQ. ID. NO. 188) are eachsoluble at greater than 3 mg/ml at a pH in the pH range of pH 6 to pH 8.

The pH-stability profiles for each peptide DPI-21.2 (SEQ. ID. NO. 28),DFI-22.2 (SEQ. ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165), DFI-26.6 (SEQ.ID. NO. 168), DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14 (SEQ. ID. NO.180) and DPII-25.15 (SEQ. ID. NO. 188) in equal concentrationcombination at 3, 2, and 1 mg/ml per peptide and prepared in duplicateas described above with one set stored at about 5° C. for 24 hours andone set stored at about 22° C.±2 for 24 hours, were calculated andtabulated as a function of the percentage of total of degradant peakarea (FIG. 38). As can be seen from the data in FIG. 38, the percentdegradation of peptide at each concentration was less for peptidesstored at about 5° C. for 24 hours as compared to peptides stored at 22°C.±2 for 24 hours over the critical pH range of pH 6.0 to pH 8.0.However, acceptable solution stability is demonstrated for all thepeptides in a common "window" within the pH range of pH 6 to pH 8 ateither temperature.

Therefore, each of peptides DPI-21.2 (SEQ. ID. NO. 28), DFI-22.2 (SEQ.ID. NO. 93), DFI-23.31(SEQ. ID. NO. 165), DFI-26.6 (SEQ. ID. NO. 168),DPII-20.9 (SEQ. ID. NO. 169), DPII-22.14 (SEQ. ID. NO. 180) andDPII-25.15 (SEQ. ID. NO. 188) was determined to possess the appropriatesolubility and stability required of a unique peptide of the invention.

Although the invention has been described with reference to itspreferred embodiments, other embodiments can achieve the same results.Variations and modifications to the present invention will be obvious tothose skilled in the art and is intended to cover in the appended claimsall such modifications and equivalents that follow in the true spiritand scope of the invention.

    __________________________________________________________________________    SEQUENCE LISTING                                                              (1) GENERAL INFORMATION:                                                      (iii) NUMBER OF SEQUENCES: 207                                                (2) INFORMATION FOR SEQ ID NO:1:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 834 base pairs                                                    (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA                                                      (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 1..738                                                          (xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:                                       AAAAACCGATTTTTGATGAGTGCAGAAGCTTTTGAACACCTCAAAACT48                            LysAsnArgPheLeuMetSerAlaGluAlaPheGluHisLeuLysThr                              151015                                                                        CAATTCGATTTGAATGCTGAAACTAACGCCTGCAGTATCAATGGAAAT96                            GlnPheAspLeuAsnAlaGluThrAsnAlaCysSerIleAsnGlyAsn                              202530                                                                        GCTCCAGCTGAAATCGATTTGCGACAAATGCGAACTGTCACTCCCATT144                           AlaProAlaGluIleAspLeuArgGlnMetArgThrValThrProIle                              354045                                                                        CGTATGCAAGGAGGCTGTGGTTCATGTTGGGCTTTCTCTGGTGTTGCC192                           ArgMetGlnGlyGlyCysGlySerCysTrpAlaPheSerGlyValAla                              505560                                                                        GCAACTGAATCAGCTTATTTGGCTCACCGTAATCAATCATTGGATCTT240                           AlaThrGluSerAlaTyrLeuAlaHisArgAsnGlnSerLeuAspLeu                              65707580                                                                      GCTGAACAAGAATTAGTCGATTGTGCTTCCCAACACGGTTGTCATGGT288                           AlaGluGlnGluLeuValAspCysAlaSerGlnHisGlyCysHisGly                              859095                                                                        GATACCATTCCACGTGGTATTGAATACATCCAACATAATGGTGTCGTC336                           AspThrIleProArgGlyIleGluTyrIleGlnHisAsnGlyValVal                              100105110                                                                     CAAGAAAGCTACTATCGATACGTTGCACGAGAACAATCATGCCGACGA384                           GlnGluSerTyrTyrArgTyrValAlaArgGluGlnSerCysArgArg                              115120125                                                                     CCAAATGCACAACGTTTCGGTATCTCAAACTATTGCCAAATTTACCCA432                           ProAsnAlaGlnArgPheGlyIleSerAsnTyrCysGlnIleTyrPro                              130135140                                                                     CCAAATGCAAACAAAATTCGTGAAGCTTTGGCTCAAACCCACAGCGCT480                           ProAsnAlaAsnLysIleArgGluAlaLeuAlaGlnThrHisSerAla                              145150155160                                                                  ATTGCCGTCATTATTGGCATCAAAGATTTAGACGCATTCCGTCATTAT528                           IleAlaValIleIleGlyIleLysAspLeuAspAlaPheArgHisTyr                              165170175                                                                     GATGGCCGAACAATCATTCAACGCGATAATGGTTACCAACCAAACTAT576                           AspGlyArgThrIleIleGlnArgAspAsnGlyTyrGlnProAsnTyr                              180185190                                                                     CACGCTGTCAACATTGTTGGTTACAGTAACGCACAAGGTGTCGATTAT624                           HisAlaValAsnIleValGlyTyrSerAsnAlaGlnGlyValAspTyr                              195200205                                                                     TGGATCGTACGAAACAGTTGGGATACCAATTGGGGTGATAATGGTTAC672                           TrpIleValArgAsnSerTrpAspThrAsnTrpGlyAspAsnGlyTyr                              210215220                                                                     GGTTATTTTGCTGCCAACATCGATTTGATGATGATTGAAGAATATCCA720                           GlyTyrPheAlaAlaAsnIleAspLeuMetMetIleGluGluTyrPro                              225230235240                                                                  TATGTTGTCATTCTCTAAACAAAAAGACAATTTCTTATATGATTGTCACTAATTT775                    TyrValValIleLeu                                                               245                                                                           ATTTAAAATCAAAATTTTTTAGAAAATGAATAAATTCATTCACAAAAATTAAAAAAAAA834                (2) INFORMATION FOR SEQ ID NO:2:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 245 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:                                       LysAsnArgPheLeuMetSerAlaGluAlaPheGluHisLeuLysThr                              151015                                                                        GlnPheAspLeuAsnAlaGluThrAsnAlaCysSerIleAsnGlyAsn                              202530                                                                        AlaProAlaGluIleAspLeuArgGlnMetArgThrValThrProIle                              354045                                                                        ArgMetGlnGlyGlyCysGlySerCysTrpAlaPheSerGlyValAla                              505560                                                                        AlaThrGluSerAlaTyrLeuAlaHisArgAsnGlnSerLeuAspLeu                              65707580                                                                      AlaGluGlnGluLeuValAspCysAlaSerGlnHisGlyCysHisGly                              859095                                                                        AspThrIleProArgGlyIleGluTyrIleGlnHisAsnGlyValVal                              100105110                                                                     GlnGluSerTyrTyrArgTyrValAlaArgGluGlnSerCysArgArg                              115120125                                                                     ProAsnAlaGlnArgPheGlyIleSerAsnTyrCysGlnIleTyrPro                              130135140                                                                     ProAsnAlaAsnLysIleArgGluAlaLeuAlaGlnThrHisSerAla                              145150155160                                                                  IleAlaValIleIleGlyIleLysAspLeuAspAlaPheArgHisTyr                              165170175                                                                     AspGlyArgThrIleIleGlnArgAspAsnGlyTyrGlnProAsnTyr                              180185190                                                                     HisAlaValAsnIleValGlyTyrSerAsnAlaGlnGlyValAspTyr                              195200205                                                                     TrpIleValArgAsnSerTrpAspThrAsnTrpGlyAspAsnGlyTyr                              210215220                                                                     GlyTyrPheAlaAlaAsnIleAspLeuMetMetIleGluGluTyrPro                              225230235240                                                                  TyrValValIleLeu                                                               245                                                                           (2) INFORMATION FOR SEQ ID NO:3:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 588 base pairs                                                    (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA                                                      (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 69..509                                                         (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3:                                       CACAAATTCTTCTTTCTTCCTTACTACTGATCATTAATCTGAAAACAAAACCAAACAAAC60                CATTCAAAATGATGTACAAAATTTTGTGTCTTTCATTGTTGGTCGCAGCC110                         MetMetTyrLysIleLeuCysLeuSerLeuLeuValAlaAla                                    1510                                                                          GTTGCTCGTGATCAAGTCGATGTCAAAGATTGTGCCAATCATGAAATC158                           ValAlaArgAspGlnValAspValLysAspCysAlaAsnHisGluIle                              15202530                                                                      AAAAAAGTTTTGGTACCAGGATGCCATGGTTCAGAACCATGTATCATT206                           LysLysValLeuValProGlyCysHisGlySerGluProCysIleIle                              354045                                                                        CATCGTGGTAAACCATTCCAATTGGAAGCCGTTTTCGAAGCCAACCAA254                           HisArgGlyLysProPheGlnLeuGluAlaValPheGluAlaAsnGln                              505560                                                                        AACACAAAAACGGCTAAAATTGAAATCAAAGCCTCAATCGATGGTTTA302                           AsnThrLysThrAlaLysIleGluIleLysAlaSerIleAspGlyLeu                              657075                                                                        GAAGTTGATGTTCCCGGTATCGATCCAAATGCATGCCATTACATGAAA350                           GluValAspValProGlyIleAspProAsnAlaCysHisTyrMetLys                              808590                                                                        TGCCCATTGGTTAAAGGACAACAATATGATATTAAATATACATGGAAT398                           CysProLeuValLysGlyGlnGlnTyrAspIleLysTyrThrTrpAsn                              95100105110                                                                   GTTCCGAAAATTGCACCAAAATCTGAAAATGTTGTCGTCACTGTTAAA446                           ValProLysIleAlaProLysSerGluAsnValValValThrValLys                              115120125                                                                     GTTATGGGTGATGATGGTGTTTTGGCCTGTGCTATTGCTACTCATGCT494                           ValMetGlyAspAspGlyValLeuAlaCysAlaIleAlaThrHisAla                              130135140                                                                     AAAATCCGCGATTAAATAAACAAAATTTATTGATTTTGTAATCACAAATGAT546                       LysIleArgAsp                                                                  145                                                                           TGATTTTCTTTCCAAAAAAAAAATAAATAAAATTTTGGGAAT588                                 (2) INFORMATION FOR SEQ ID NO:4:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 146 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4:                                       MetMetTyrLysIleLeuCysLeuSerLeuLeuValAlaAlaValAla                              151015                                                                        ArgAspGlnValAspValLysAspCysAlaAsnHisGluIleLysLys                              202530                                                                        ValLeuValProGlyCysHisGlySerGluProCysIleIleHisArg                              354045                                                                        GlyLysProPheGlnLeuGluAlaValPheGluAlaAsnGlnAsnThr                              505560                                                                        LysThrAlaLysIleGluIleLysAlaSerIleAspGlyLeuGluVal                              65707580                                                                      AspValProGlyIleAspProAsnAlaCysHisTyrMetLysCysPro                              859095                                                                        LeuValLysGlyGlnGlnTyrAspIleLysTyrThrTrpAsnValPro                              100105110                                                                     LysIleAlaProLysSerGluAsnValValValThrValLysValMet                              115120125                                                                     GlyAspAspGlyValLeuAlaCysAlaIleAlaThrHisAlaLysIle                              130135140                                                                     ArgAsp                                                                        145                                                                           (2) INFORMATION FOR SEQ ID NO:5:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 1072 base pairs                                                   (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA                                                      (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 36..1001                                                        (xi) SEQUENCE DESCRIPTION: SEQ ID NO:5:                                       CGTTTTCTTCCATCAAAATTAAAAATTCATCAAAAATGAAATTCGTTTTGGCC53                       MetLysPheValLeuAla                                                            15                                                                            ATTGCCTCTTTGTTGGTATTGAGCACTGTTTATGCTCGTCCAGCTTCA101                           IleAlaSerLeuLeuValLeuSerThrValTyrAlaArgProAlaSer                              101520                                                                        ATCAAAACTTTTGAAGAATTCAAAAAAGCCTTCAACAAAAACTATGCC149                           IleLysThrPheGluGluPheLysLysAlaPheAsnLysAsnTyrAla                              253035                                                                        ACCGTTGAAGAGGAAGAAGTTGCCCGTAAAAACTTTTTGGAATCATTG197                           ThrValGluGluGluGluValAlaArgLysAsnPheLeuGluSerLeu                              404550                                                                        AAATATGTTGAAGCTAACAAAGGTGCCATCAACCATTTGTCCGATTTG245                           LysTyrValGluAlaAsnLysGlyAlaIleAsnHisLeuSerAspLeu                              55606570                                                                      TCATTGGATGAATTCAAAAACCGTTATTTGATGAGTGCTGAAGCTTTT293                           SerLeuAspGluPheLysAsnArgTyrLeuMetSerAlaGluAlaPhe                              758085                                                                        GAACAACTCAAAACTCAATTCGATTTGAATGCCGAAACAAGCGCTTGC341                           GluGlnLeuLysThrGlnPheAspLeuAsnAlaGluThrSerAlaCys                              9095100                                                                       CGTATCAATTCGGTTAACGTTCCATCGGAATTGGATTTACGATCACTG389                           ArgIleAsnSerValAsnValProSerGluLeuAspLeuArgSerLeu                              105110115                                                                     CGAACTGTCACTCCAATCCGTATGCAAGGAGGCTGTGGTTCATGTTGG437                           ArgThrValThrProIleArgMetGlnGlyGlyCysGlySerCysTrp                              120125130                                                                     GCTTTCTCTGGTGTTGCCGCAACTGAATCAGCTTATTTGGCCTACCGT485                           AlaPheSerGlyValAlaAlaThrGluSerAlaTyrLeuAlaTyrArg                              135140145150                                                                  AACACGTCTTTGGATCTTTCTGAACAGGAACTCGTCGATTGCGCATCT533                           AsnThrSerLeuAspLeuSerGluGlnGluLeuValAspCysAlaSer                              155160165                                                                     CAACACGGATGTCACGGCGATACAATACCAAGAGGCATCGAATACATC581                           GlnHisGlyCysHisGlyAspThrIleProArgGlyIleGluTyrIle                              170175180                                                                     CAACAAAATGGTGTCGTTGAAGAAAGAAGCTATCCATACGTTGCACGA629                           GlnGlnAsnGlyValValGluGluArgSerTyrProTyrValAlaArg                              185190195                                                                     GAACAACGATGCCGACGACCAAATTCGCAACATTACGGTATCTCAAAC677                           GluGlnArgCysArgArgProAsnSerGlnHisTyrGlyIleSerAsn                              200205210                                                                     TACTGCCAAATTTATCCACCAGATGTGAAACAAATCCGTGAAGCTTTG725                           TyrCysGlnIleTyrProProAspValLysGlnIleArgGluAlaLeu                              215220225230                                                                  ACTCAAACACACACAGCTATTGCCGTCATTATTGGCATCAAAGATTTG773                           ThrGlnThrHisThrAlaIleAlaValIleIleGlyIleLysAspLeu                              235240245                                                                     AGAGCTTTCCAACATTATGATGGACGAACAATCATTCAACATGACAAT821                           ArgAlaPheGlnHisTyrAspGlyArgThrIleIleGlnHisAspAsn                              250255260                                                                     GGTTATCAACCAAACTATCATGCCGTCAACATTGTCGGTTACGGAAGT869                           GlyTyrGlnProAsnTyrHisAlaValAsnIleValGlyTyrGlySer                              265270275                                                                     ACACAAGGCGACGATTATTGGATCGTACGAAACAGTTGGGATACTACC917                           ThrGlnGlyAspAspTyrTrpIleValArgAsnSerTrpAspThrThr                              280285290                                                                     TGGGGAGATAGCGGATACGGATATTTCCAAGCCGGAAACAACCTCATG965                           TrpGlyAspSerGlyTyrGlyTyrPheGlnAlaGlyAsnAsnLeuMet                              295300305310                                                                  ATGATCGAACAATATCCATATGTTGTAATCATGTGAACATTTGAAATTGAATA1018                     MetIleGluGlnTyrProTyrValValIleMet                                             315320                                                                        TATTTATTTGTTTTCAAAATAAAAACAACTACTCTTGCGAGTATTTTTTACTCG1072                    (2) INFORMATION FOR SEQ ID NO:6:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 321 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:6:                                       MetLysPheValLeuAlaIleAlaSerLeuLeuValLeuSerThrVal                              151015                                                                        TyrAlaArgProAlaSerIleLysThrPheGluGluPheLysLysAla                              202530                                                                        PheAsnLysAsnTyrAlaThrValGluGluGluGluValAlaArgLys                              354045                                                                        AsnPheLeuGluSerLeuLysTyrValGluAlaAsnLysGlyAlaIle                              505560                                                                        AsnHisLeuSerAspLeuSerLeuAspGluPheLysAsnArgTyrLeu                              65707580                                                                      MetSerAlaGluAlaPheGluGlnLeuLysThrGlnPheAspLeuAsn                              859095                                                                        AlaGluThrSerAlaCysArgIleAsnSerValAsnValProSerGlu                              100105110                                                                     LeuAspLeuArgSerLeuArgThrValThrProIleArgMetGlnGly                              115120125                                                                     GlyCysGlySerCysTrpAlaPheSerGlyValAlaAlaThrGluSer                              130135140                                                                     AlaTyrLeuAlaTyrArgAsnThrSerLeuAspLeuSerGluGlnGlu                              145150155160                                                                  LeuValAspCysAlaSerGlnHisGlyCysHisGlyAspThrIlePro                              165170175                                                                     ArgGlyIleGluTyrIleGlnGlnAsnGlyValValGluGluArgSer                              180185190                                                                     TyrProTyrValAlaArgGluGlnArgCysArgArgProAsnSerGln                              195200205                                                                     HisTyrGlyIleSerAsnTyrCysGlnIleTyrProProAspValLys                              210215220                                                                     GlnIleArgGluAlaLeuThrGlnThrHisThrAlaIleAlaValIle                              225230235240                                                                  IleGlyIleLysAspLeuArgAlaPheGlnHisTyrAspGlyArgThr                              245250255                                                                     IleIleGlnHisAspAsnGlyTyrGlnProAsnTyrHisAlaValAsn                              260265270                                                                     IleValGlyTyrGlySerThrGlnGlyAspAspTyrTrpIleValArg                              275280285                                                                     AsnSerTrpAspThrThrTrpGlyAspSerGlyTyrGlyTyrPheGln                              290295300                                                                     AlaGlyAsnAsnLeuMetMetIleGluGlnTyrProTyrValValIle                              305310315320                                                                  Met                                                                           (2) INFORMATION FOR SEQ ID NO:7:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 491 base pairs                                                    (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA                                                      (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 1..390                                                          (xi) SEQUENCE DESCRIPTION: SEQ ID NO:7:                                       GATCAAGTCGATGTTAAAGATTGTGCCAACAATGAAATCAAAAAAGTA48                            AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        ATGGTCGATGGTTGCCATGGTTCTGATCCATGCATAATCCATCGTGGT96                            MetValAspGlyCysHisGlySerAspProCysIleIleHisArgGly                              202530                                                                        AAACCATTCACTTTGGAAGCCTTATTCGATGCCAACCAAAACACTAAA144                           LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        ACCGCTAAAACTGAAATCAAAGCCAGCCTCGATGGTCTTGAAATTGAT192                           ThrAlaLysThrGluIleLysAlaSerLeuAspGlyLeuGluIleAsp                              505560                                                                        GTTCCCGGTATTGATACCAATGCTTGCCATTTTATGAAATGTCCATTG240                           ValProGlyIleAspThrAsnAlaCysHisPheMetLysCysProLeu                              65707580                                                                      GTTAAAGGTCAACAATATGATGCCAAATATACATGGAATGTGCCCAAA288                           ValLysGlyGlnGlnTyrAspAlaLysTyrThrTrpAsnValProLys                              859095                                                                        ATTGCACCAAAATCTGAAAACGTTGTCGTTACAGTCAAACTTGTTGGT336                           IleAlaProLysSerGluAsnValValValThrValLysLeuValGly                              100105110                                                                     GATAATGGTGTTTTGGCTTGCGCTATTGCTACCCACGCTAAAATCCGT384                           AspAsnGlyValLeuAlaCysAlaIleAlaThrHisAlaLysIleArg                              115120125                                                                     GATTAAAAAAAAAAAATAAATATGAAAATTTTCACCAACATCGAACAAAATTC437                      Asp                                                                           130                                                                           AATAACCAAAATTTGAATCAAAAACGGAATTCCAAGCTGAGCGCCGGTCGCTAC491                     (2) INFORMATION FOR SEQ ID NO:8:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 129 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:8:                                       AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        MetValAspGlyCysHisGlySerAspProCysIleIleHisArgGly                              202530                                                                        LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        ThrAlaLysThrGluIleLysAlaSerLeuAspGlyLeuGluIleAsp                              505560                                                                        ValProGlyIleAspThrAsnAlaCysHisPheMetLysCysProLeu                              65707580                                                                      ValLysGlyGlnGlnTyrAspAlaLysTyrThrTrpAsnValProLys                              859095                                                                        IleAlaProLysSerGluAsnValValValThrValLysLeuValGly                              100105110                                                                     AspAsnGlyValLeuAlaCysAlaIleAlaThrHisAlaLysIleArg                              115120125                                                                     Asp                                                                           (2) INFORMATION FOR SEQ ID NO:9:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9:                                       ThrAsnAlaCysSerIleAsnGlyAsnAlaProAlaGluIleAspLeu                              151015                                                                        ArgGlnMetArg                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:10:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:10:                                      GluIleAspLeuArgGlnMetArgThrValThrProIleArgMetGln                              151015                                                                        GlyGlyCysGlySerCysTrpAlaPheSerGly                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:11:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:11:                                      ThrValThrProIleArgMetGlnGlyGlyCysGlySerCysTrpAla                              151015                                                                        PheSerGlyValAlaAlaThrGluSerAlaTyrLeuAla                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:12:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:12:                                      ValAlaAlaThrGluSerAlaTyrLeuAlaHisArgAsnGlnSerLeu                              151015                                                                        AspLeuAlaGluGln                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:13:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:13:                                      HisArgAsnGlnSerLeuAspLeuAlaGluGlnGluLeuValAspCys                              151015                                                                        AlaSerGlnHisGlyCys                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:14:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:14:                                      GluLeuValAspCysAlaSerGlnHisGlyCysHisGlyAspThrIle                              151015                                                                        ProArgGlyIleGlu                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:15:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:15:                                      GlyAspThrIleProArgGlyIleGluTyrIleGlnHisAsnGlyVal                              151015                                                                        ValGlnGluSerTyrTyrArgTyrValAlaArgGlu                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:16:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:16:                                      GluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyr                              151015                                                                        ValAlaArgGlu                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:17:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:17:                                      HisAsnGlyValValGlnGluSerTyrTyrArgTyrValAlaArgGlu                              151015                                                                        GlnSerCysArgArgProAsnAlaGln                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:18:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 19 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:18:                                      GlnSerCysArgArgProAsnAlaGlnArgPheGlyIleSerAsnTyr                              151015                                                                        CysGlnIle                                                                     (2) INFORMATION FOR SEQ ID NO:19:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:19:                                      ArgPheGlyIleSerAsnTyrCysGlnIleTyrProProAsnAlaAsn                              151015                                                                        LysIleArgGluAlaLeu                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:20:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:20:                                      TyrProProAsnAlaAsnLysIleArgGluAlaLeuAlaGlnThrHis                              151015                                                                        SerAlaIleAlaValIleIleGly                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:21:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:21:                                      AlaGlnThrHisSerAlaIleAlaValIleIleGlyIleLysAspLeu                              151015                                                                        AspAlaPheArgHisTyrAspGlyArgThr                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:22:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:22:                                      IleLysAspLeuAspAlaPheArgHisTyrAspGlyThrArgIleIle                              151015                                                                        GlnArgAspAsnGlyTyrGlnProAsnTyr                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:23:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:23:                                      IleIleGlnArgAspAsnGlyTyrGlnProAsnTyrHisAlaValAsn                              151015                                                                        IleValGlyTyrSerAsnAla                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:24:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:24:                                      HisAlaValAsnIleValGlyTyrSerAsnAlaGlnGlyValAspTyr                              151015                                                                        TrpIleValArgAsnSer                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:25:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:25:                                      GlnGlyValAspTyrTrpIleValArgAsnSerTrpAspThrAsnTrp                              151015                                                                        GlyAspAsnGlyTyrGlyTyrPhe                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:26:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:26:                                      GlyAspAsnGlyTyrGlyTyrPheAlaAlaAsnIleAspLeuMetMet                              151015                                                                        IleGluGluTyrProTyrValValIleLeu                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:27:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:27:                                      ThrAsnAlaCysSerIleAsnGlyAsnAlaProAlaGluIleAspLeu                              151015                                                                        ArgGlnMetArgThrValThrProIleArgMetGln                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:28:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:                                      SerIleAsnGlyAsnAlaProAlaGluIleAspLeuArgGlnMetArg                              151015                                                                        ThrValThrProIleArgMetGln                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:29:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:29:                                      AlaPheSerGlyValAlaAlaThrGluSerAlaTyrLeuAlaHisArg                              151015                                                                        AsnGlnSerLeuAspLeuAlaGluGlnGluLeuValAsp                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:30:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:30:                                      ValAlaAlaThrGluSerAlaTyrLeuAlaHisArgAsnGlnSerLeu                              151015                                                                        AspLeuAlaGluGlnGluLeuValAsp                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:31:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:31:                                      AlaPheSerGlyValAlaAlaThrGluSerAlaTyrLeuAlaHisArg                              151015                                                                        AsnGlnSerLeuAspLeuAlaGluGln                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:32:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:32:                                      ValAlaAlaThrGluSerAlaTyrLeuAlaHisArgAsnGlnSerLeu                              151015                                                                        AspLeuAlaGluGlnGluLeuValAspCysAlaSerGln                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:33:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:33:                                      GluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyr                              151015                                                                        ValAlaArgGluGlnCysArgArgProAsnAlaGln                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:34:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:34:                                      AspThrIleProArgGlyIleGluTyrIleGlnHisAsnGlyValVal                              151015                                                                        GlnGluSerTyrTyrArgTyrValAlaArgGluGlnSer                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:35:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:35:                                      GlnIleTyrProProAsnAlaAsnLysIleArgGluAlaLeuAlaGln                              151015                                                                        ThrHisSerAlaIleAlaValIleIleGlyIleLysAsp                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:36:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:36:                                      GlnIleTyrProProAsnAlaAsnLysIleArgGluAlaLeuAlaGln                              151015                                                                        ThrHisSerAlaIleAla                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:37:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:37:                                      IleIleGlyIleLysAspLeuAspAlaPheArgHisTyrAspGlyArg                              151015                                                                        ThrIleIleGlnArgAspAsnGlyTyrGln                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:38:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:38:                                      ArgAspAsnGlyTyrGlnPheAsnTyrHisAlaValAsnIleValGly                              151015                                                                        TyrSerAsnAlaGlnGlyValAspTyr                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:39:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:39:                                      AsnIleValGlyTyrSerAsnAlaGlnGlyValAspTyrTrpIleVal                              151015                                                                        ArgAsnSerTrpAspThrAsnTrpGlyAspAsnGlyTyr                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:40:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:40:                                      AsnIleValGlyTyrSerAsnAlaGlnGlyValAspTyrTrpIleVal                              151015                                                                        ArgAsnSerTrpAspThrAsn                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:41:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:41:                                      AspGlnValAspValLysAspCysAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGly                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:42:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:42:                                      HisGluIleLysLysValLeuValProGlyCysHisGlySerGluPro                              151015                                                                        CysIleIleHisArgGlyLysProPhe                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:43:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:43:                                      HisGlySerGluProCysIleIleHisArgGlyLysProPheGlnLeu                              151015                                                                        GluAlaValPheGluAlaAsnGlnAsnThrLysThrAla                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:44:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:44:                                      GlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThrAlaLys                              151015                                                                        IleGluIleLysAlaSerIleAspGly                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:45:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:45:                                      LysIleGluIleLysAlaSerIleAspGlyLeuGluValAspValPro                              151015                                                                        GlyIleAspProAsnAlaCysHisTyrMetLys                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:46:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:46:                                      LeuGluValAspValProGlyIleAspProAsnAlaCysHisTyrMet                              151015                                                                        LysCysProLeuValLysGlyGlnGlnTyr                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:47:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:47:                                      CysProLeuValLysGlyGlnGlnTyrAspIleLysTyrThrTrpAsn                              151015                                                                        ValProLysIleAlaProLysSerGluAsnVal                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:48:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:48:                                      AspIleLysTyrThrTrpAsnValProLysIleAlaProLysSerGlu                              151015                                                                        AsnValValValThrValLysValMetGly                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:49:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:49:                                      ValValThrValLysValMetGlyAspAspGlyValLeuAlaCysAla                              151015                                                                        IleAlaThrHisAlaLysIleArgAsp                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:50:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:50:                                      AspGlnValAspValLysAspCysAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyCysHisGlySerGluPro                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:51:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:51:                                      AspGlnValAspValLysAspGluAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyCysHisGlySerGluPro                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:52:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:52:                                      AspGlnValAspValLysAspSerAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyCysHisGlySerGluPro                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:53:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:53:                                      AspGlnValAspValLysAspCysAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyGluHisGlySerGluPro                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:54:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:54:                                      AspGlnValAspValLysAspCysAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlySerHisGlySerGluPro                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:55:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:55:                                      AspGlnValAspValLysAspGluAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyGluHisGlySerGluPro                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:56:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:56:                                      AspGlnValAspValLysAspSerAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlySerHisGlySerGluPro                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:57:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:57:                                      AspGlnValAspValLysAspGluAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGly                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:58:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:58:                                      AspGlnValAspValLysAspSerAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGly                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:59:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 16 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:59:                                      HisGluIleLysLysValLeuValProGlyCysHisGlySerGluPro                              151015                                                                        (2) INFORMATION FOR SEQ ID NO:60:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 16 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:60:                                      HisGluIleLysLysValLeuValProGlyGluHisGlySerGluPro                              151015                                                                        (2) INFORMATION FOR SEQ ID NO:61:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 16 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:61:                                      HisGluIleLysLysValLeuValProGlySerHisGlySerGluPro                              151015                                                                        (2) INFORMATION FOR SEQ ID NO:62:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:62:                                      LysProPheGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLys                              151015                                                                        ThrAlaLysIleGluIleLysAlaSerThrAspGly                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:63:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:63:                                      GlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThrAlaLys                              151015                                                                        IleGluIleLysAlaSerIleAspGlyLeuGluVal                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:64:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:64:                                      PheGluAlaAsnGlnAsnThrLysThrAlaLysIleGluIleLysAla                              151015                                                                        SerIleAspGlyLeuGluValAspValProGly                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:65:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:65:                                      GlnAsnThrLysThrAlaLysIleGluIleLysAlaSerIleAspGly                              151015                                                                        LeuGluValAspValProGly                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:66:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:66:                                      ProLeuValLysGlyGlnGlnTyrAspIleLysTyrThrTrpAsnVal                              151015                                                                        ProLysIleAlaProLysSerGluAsnVal                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:67:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:67:                                      ProLeuValLysGlyGlnGlnTyrAspIleLysTyrThrTyrAsnVal                              151015                                                                        ProLysIleAlaProLysSerGluAsnVal                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:68:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 13 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:68:                                      LysSerGluAsnValValValThrValLysValMetGly                                       1510                                                                          (2) INFORMATION FOR SEQ ID NO:69:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:69:                                      ThrValLysValMetGlyAspAspGlyValLeuAlaCysAlaIleAla                              151015                                                                        ThrHisAlaLysIleArgAsp                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:70:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:70:                                      ThrValLysValMetGlyAspAspGlyValLeuAlaGluAlaIleAla                              151015                                                                        ThrHisAlaLysIleArgAsp                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:71:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:71:                                      ThrValLysValLeuGlyAspAspGlyValLeuAlaSerAlaIleAla                              151015                                                                        ThrHisAlaLysIleArgAsp                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:72:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:72:                                      ThrSerAlaCysArgIleAsnSerValAsnValProSerGluLeuAsp                              151015                                                                        LeuArgSerLeuArg                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:73:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:73:                                      GluLeuAspLeuArgSerLeuArgThrValThrProIleArgMetGln                              151015                                                                        GlyGlyCysGlySerCysTrpAlaPheSerGly                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:74:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:74:                                      ThrValThrProIleArgMetGlnGlyGlyCysGlySerCysTrpAla                              151015                                                                        PheSerGlyValAlaAlaThrGluSerAlaTyrLeuAla                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:75:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:75:                                      ValAlaAlaThrGluSerAlaTyrLeuAlaTyrArgAsnThrSerLeu                              151015                                                                        AspLeuSerGluGln                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:76:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:76:                                      TyrArgAsnThrSerLeuAspLeuSerGluGlnGluLeuValAspCys                              151015                                                                        AlaSerGlnHisGlyCys                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:77:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:77:                                      GluLeuValAspCysAlaSerGlnHisGlyCysHisGlyAspThrIle                              151015                                                                        ProArgGlyIleGlu                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:78:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:78:                                      GlyAspThrIleProArgGlyIleGluTyrIleGlnGlnAsnGlyVal                              151015                                                                        ValGluGluArgSerTyrProTyrValAlaArgGlu                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:79:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:79:                                      GlnAsnGlyValValGluGluArgSerTyrProTyrValAlaArgGlu                              151015                                                                        GlnArgCysArgArgProAsnSerGln                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:80:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 19 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:80:                                      GlnArgCysArgArgProAsnSerGlnHisTyrGlyIleSerAsnTyr                              151015                                                                        CysGlnIle                                                                     (2) INFORMATION FOR SEQ ID NO:81:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:81:                                      HisTyrGlyIleSerAsnTyrCysGlnIleTyrProProAspValLys                              151015                                                                        GlnIleArgGluAlaLeu                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:82:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:82:                                      TyrProProAspValLysGlnIleArgGluAlaLeuPheGlnThrHis                              151015                                                                        ThrAlaIleAlaValIleIleGly                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:83:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:83:                                      ThrGlnThrHisThrAlaIleAlaValIleIleGlyIleLysAspLeu                              151015                                                                        ArgAlaPheGlnHisTyrAspGlyArgThr                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:84:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:84:                                      IleLysAspLeuArgAlaPheGlnHisTyrAspGlyArgThrIleIle                              151015                                                                        GlnHisAspAsnGlyTyrGlnProAsnTyr                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:85:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:85:                                      AspGlyArgThrIleIleGlnHisAspAsnGlyTyrGlnProAsn                                 151015                                                                        (2) INFORMATION FOR SEQ ID NO:86:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:86:                                      IleIleGlnHisAspAsnGlyTyrGlnProAsnTyrHisAlaValAsn                              151015                                                                        IleValGlyTyrGlySerThr                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:87:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:87:                                      HisAlaValAsnIleValGlyTyrGlySerThrGlnGlyAspAspTyr                              151015                                                                        TrpIleValArgAsnSer                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:88:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:88:                                      GlnGlyAspAspTyrTrpIleValArgAsnSerTrpAspThrThrTrp                              151015                                                                        GlyAspSerGlyTyrGlyTyrPhe                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:89:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:89:                                      GlyAspSerGlyTyrGlyTyrPheGlnAlaGlyAsnAsnLeuMetMet                              151015                                                                        IleGluGlnTyrProTyrValValIleMet                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:90:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:90:                                      ThrSerAlaCysArgIleAsnSerValAsnValProSerGluLeuAsp                              151015                                                                        LeuArgSerLeuArgThrValThrProIleArgMetGln                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:91:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:91:                                      ArgIleAsnSerValAsnValProSerGluLeuAspLeuArgSerLeu                              151015                                                                        ArgThrValThrProIleArgMetGln                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:92:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:92:                                      AlaPheSerGlyValAlaAlaThrGluSerAlaTyrLeuAlaTyrArg                              151015                                                                        AsnThrSerLeuAspLeuSerGluGlnGluLeuValAsp                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:93:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:93:                                      ValAlaAlaThrGluSerAlaTyrLeuAlaTyrArgAsnThrSerLeu                              151015                                                                        AspLeuSerGluGlnGluLeuValAsp                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:94:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:94:                                      ValAlaAlaThrGluSerAlaTyrLeuAlaTyrArgAsnThrSerLeu                              151015                                                                        AspLeuSerGluGlnGluLeuValAspCysAlaSerGln                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:95:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:95:                                      GluTyrIleGlnGlnAsnGlyValValGluGluArgSerTyrProTyr                              151015                                                                        ValAlaArgGluGlnArgCysArgArgProAsnSerGln                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:96:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:96:                                      AspThrIleProArgGlyIleGluTyrIleGlnGlnAsnGlyValVal                              151015                                                                        GluGluArgSerTyrProTyrValAlaArgGluGlnArg                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:97:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:97:                                      GlnIleTyrProProAspValLysGlnIleArgGluAlaLeuThrGln                              151015                                                                        ThrHisThrAlaIleAlaValIleIleGlyIleLysAsp                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:98:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:98:                                      GlnIleTyrProProAspValLysGlnIleArgGluAlaLeuThrGln                              151015                                                                        ThrHisThrAlaIleAla                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:99:                                             (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:99:                                      IleIleGlyIleLysAspLeuArgAlaPheGlnHisTyrAspGlyArg                              151015                                                                        ThrIleIleGlnHisAspAsnGlyTyrGln                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:100:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:100:                                     HisAspAsnGlyTyrGlnProAsnTyrHisAlaValAsnIleValGly                              151015                                                                        TyrGlySerThrGlnGlyAspAspTyr                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:101:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:101:                                     AsnIleValGlyTyrGlySerThrGlnGlyAspAspTyrTrpIleVal                              151015                                                                        ArgAsnSerTrpAspThrThrTrpGlyAspSerGlyTyr                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:102:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:102:                                     AsnIleValGlyTyrGlySerThrGlnGlyAspAspTyrTrpIleVal                              151015                                                                        ArgAsnSerTrpAspThrThr                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:103:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:103:                                     AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        MetValAspGly                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:104:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:104:                                     AsnGluIleLysLysValMetValAspGlyCysHisGlySerAspPro                              151015                                                                        CysIleIleHisArgGlyLysProPhe                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:105:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:105:                                     HisGlySerAspProCysIleIleHisArgGlyLysProPheThrLeu                              151015                                                                        GluAlaLeuPheAspAlaAsnGlnAsnThrLysThrAla                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:106:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 19 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:106:                                     HisArgGlyLysProPheThrLeuGluAlaLeuPheAspAlaAsnGln                              151015                                                                        AsnThrLys                                                                     (2) INFORMATION FOR SEQ ID NO:107:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:107:                                     ThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLysThrAlaLys                              151015                                                                        IleGluIleLysAlaSerLeuAspGly                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:108:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:108:                                     GlnAsnThrLysThrAlaLysIleGluIleLysAlaSerLeuAspGly                              151015                                                                        LeuGluIleAspValProGlyIleAspThr                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:109:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:109:                                     LysIleGluIleLysAlaSerLeuAspGlyLeuGluIleAspValPro                              151015                                                                        GlyIleAspThrAsnAlaCysHisPheMetLys                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:110:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:110:                                     LeuGluIleAspValProGlyIleAspThrAsnAlaCysHisPheMet                              151015                                                                        LysCysProLeuValLysGlyGlnGlnTyr                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:111:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:111:                                     CysProLeuValLysGlyGlnGlnTyrAspAlaLysTyrThrTrpAsn                              151015                                                                        ValProLysIleAlaProLysSerGluAsnVal                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:112:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:112:                                     AspAlaLysTyrThrTrpAsnValProLysIleAlaProLysSerGlu                              151015                                                                        AsnValValValThrValLysLeuValGly                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:113:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 35 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:113:                                     ProLysIleAlaProLysSerGluAsnValValValThrValLysLeu                              151015                                                                        ValGlyAspAsnGlyValLeuAlaCysAlaIleAlaThrHisAlaLys                              202530                                                                        IleArgAsp                                                                     35                                                                            (2) INFORMATION FOR SEQ ID NO:114:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:114:                                     ValValThrValLysLeuValGlyAspAsnGlyValLeuAlaCysAla                              151015                                                                        IleAlaThrHisAlaLysIleArgAsp                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:115:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:115:                                     LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              151015                                                                        ThrAlaLysIleGluIleLysAlaSerLeuAspGly                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:116:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:116:                                     ThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLysThrAlaLys                              151015                                                                        IleGluIleLysAlaSerLeuAspGlyLeuGluIle                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:117:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:117:                                     HisArgAsnGlnSerLeuAspLeuAlaGluGlnAspLeuValAspCys                              151015                                                                        AlaSerGlnHisGlyCys                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:118:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:118:                                     AspLeuValAspCysAlaSerGlnHisGlyCysHisGlyAspThrIle                              151015                                                                        ProArgGlyIleGlu                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:119:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: N-terminal                                                 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:119:                                     HisGlySerGluProCysIleIleHisArgGlyLysProPheGlnLeu                              151015                                                                        GluAlaValPheGluAlaValGlnAsnThrLysThrAla                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO: 120:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 120:                                    LysSerIleAsnGlyAsnAlaProAlaGluIleAspLeuArgGlnLeu                              151015                                                                        ArgThrValThrProIleArgLeuGln                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO: 121:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 31 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 121:                                    LysLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerCysArgArgProAsnAlaGln                                 202530                                                                        (2) INFORMATION FOR SEQ ID NO: 122:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 31 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 122:                                    LysLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerSerArgArgProAsnAlaGln                                 202530                                                                        (2) INFORMATION FOR SEQ ID NO: 123:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 31 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 123:                                    LysLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerGluArgArgProAsnAlaGln                                 202530                                                                        (2) INFORMATION FOR SEQ ID NO: 124:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 32 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 124:                                    AspLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerSerArgArgProAsnAlaGlnArg                              202530                                                                        (2) INFORMATION FOR SEQ ID NO: 125:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 32 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 125:                                    AspLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerCysArgArgProAsnAlaGlnArg                              202530                                                                        (2) INFORMATION FOR SEQ ID NO: 126:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 126:                                    AspGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArg                              151015                                                                        TyrValAlaArgGluGlnSerCysArgArgAspAsp                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO: 127:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 127:                                    AspGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArg                              151015                                                                        TyrValAlaArgGluGlnSerSerArgArgAspAsp                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO: 128:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 128:                                    AspGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArg                              151015                                                                        TyrValAlaArgGluGlnSerCysArgArgProAsp                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO: 129:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 129:                                    AspGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArg                              151015                                                                        TyrValAlaArgGluGlnSerSerArgArgProAsp                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO: 130:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 130:                                    AspGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArg                              151015                                                                        TyrValAlaArgGluGlnSerGlu                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO: 131:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 131:                                    ArgTyrValAlaArgGluGlnSerCysArgArgProAsnAlaGln                                 151015                                                                        (2) INFORMATION FOR SEQ ID NO: 132:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 132:                                    ArgTyrValAlaArgGluGlnSerGluArgArgProAsnAlaGln                                 151015                                                                        (2) INFORMATION FOR SEQ ID NO: 133:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 133:                                    ArgTyrValAlaArgGluGlnSerSerArgArgProAsnAlaGln                                 151015                                                                        (2) INFORMATION FOR SEQ ID NO: 134:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 134:                                    AspGluGlyIleLysAspLeuAspAlaPheArgHisTyrAspGlyArg                              151015                                                                        ThrIleIleGlnArgAspAsnGlyTyrGln                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO: 135:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 13 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 135:                                    IleGlyIleLysAspLeuAspAlaPheArgHisTyrAsp                                       1510                                                                          (2) INFORMATION FOR SEQ ID NO: 136:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 13 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 136:                                    AspAlaPheArgHisTyrAspGlyArgThrIleIleGln                                       1510                                                                          (2) INFORMATION FOR SEQ ID NO: 137:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 13 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 137:                                    ThrIleIleGlnArgAspAsnGlyTyrGlnProAsnTyr                                       1510                                                                          (2) INFORMATION FOR SEQ ID NO: 138:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 138:                                    AspGlnValAspValLysAspSerAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlySerHisGlySerGluProLys                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO: 139:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 31 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 139:                                    AspLysGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaSerIleAspGlyLeuGluValAsp                                 202530                                                                        (2) INFORMATION FOR SEQ ID NO: 140:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 31 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 140:                                    AspLysGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaSerIleAspGlyLeuGluValLys                                 202530                                                                        (2) INFORMATION FOR SEQ ID NO: 141:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 141:                                    GlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThrAlaLys                              151015                                                                        IleGluIleLysAlaSerIleAspGlu                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO: 142:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 142:                                    AspLysGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaSerIleAspGlu                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO: 143:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 13 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 143:                                    AlaLysIleGluIleLysAlaSerIleAspGlyLeuGlu                                       1510                                                                          (2) INFORMATION FOR SEQ ID NO: 144:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 144:                                    AspLysGluGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLys                              151015                                                                        ThrAlaLysIleGluIleLysAlaSerIleAspGlu                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO: 145:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 145:                                    AspLysGluGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLys                              151015                                                                        ThrAlaLysIleGluIleLysAlaSerIleAspGluGlu                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO: 146:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 146:                                    LeuGluAlaValPheGluAlaAsnGlnAsnThrLysThrAlaLys                                 151015                                                                        (2) INFORMATION FOR SEQ ID NO: 147:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 147:                                    LeuGluAlaValPheGluAlaAsnGlnAlaThrLysThrAlaLys                                 151015                                                                        (2) INFORMATION FOR SEQ ID NO: 148:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 148:                                    LysThrValLysValLeuGlyAspAspGlyValLeuAlaSerAlaIle                              151015                                                                        AlaThrHisAlaLysIleArgAsp                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO: 149:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 149:                                    AspThrValLysValLeuGlyAspAspGlyValLeuAlaSerAlaIle                              151015                                                                        AlaThrHisAlaLysIleArgAsp                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO: 150:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 10 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 150:                                    MetGlyHisHisHisHisHisHisGluPhe                                                1510                                                                          (2) INFORMATION FOR SEQ ID NO: 151:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 31 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 151:                                    LysLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerCysArgArgProAsnAlaGln                                 202530                                                                        (2) INFORMATION FOR SEQ ID NO: 152:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 30 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 152:                                    LysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArg                              151015                                                                        TyrValAlaArgGluGlnSerCysArgArgProAsnAlaGln                                    202530                                                                        (2) INFORMATION FOR SEQ ID NO: 153:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 153:                                    GluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyr                              151015                                                                        ValAlaArgGluGlnSerSerArgArgProAsnAlaGln                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO: 154:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 154:                                    GluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyr                              151015                                                                        ValAlaArgGluGlnSerGluArgArgProAsnAlaGln                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO: 155:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 30 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 155:                                    LysLysGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaSerIleAspGlyLeuGluVal                                    202530                                                                        (2) INFORMATION FOR SEQ ID NO: 156:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 30 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 156:                                    LysGlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThrAla                              151015                                                                        LysIleGluIleLysAlaSerIleAspGlyLeuGluValLys                                    202530                                                                        (2) INFORMATION FOR SEQ ID NO: 157:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 129 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 157:                                    AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        MetValProGlyCysHisGlySerGluProCysIleIleHisArgGly                              202530                                                                        LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        ThrAlaLysIleGluIleLysAlaSerLeuAspGlyLeuGluIleAsp                              505560                                                                        ValProGlyIleAspThrAsnAlaCysHisPheValLysCysProLeu                              65707580                                                                      ValLysGlyGlnGlnTyrAspIleLysTyrThrTrpAsnValProLys                              859095                                                                        IleAlaProLysSerGluAsnValValValThrValLysLeuIleGly                              100105110                                                                     AspAsnGlyValLeuAlaCysAlaIleAlaThrHisAlaLysIleArg                              115120125                                                                     Asp                                                                           (2) INFORMATION FOR SEQ ID NO: 158:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 130 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 158:                                    AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        MetValProGlyCysHisGlySerGluProCysIleIleHisArgGly                              202530                                                                        LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        ThrAlaLysIleGluIleLysAlaSerLeuAspGlyLeuGluIleAsp                              505560                                                                        ValProGlyIleAspThrAsnAlaCysHisPheMetLysCysProLeu                              65707580                                                                      ValLysGlyGlnGlnTyrAspAlaLysTyrThrTrpAsnValProLys                              859095                                                                        IleAlaProLysSerGluAsnValValValThrValLysLeuValGly                              100105110                                                                     AspAsnGlyValLeuAlaCysAlaIleIleAlaThrHisAlaLysIle                              115120125                                                                     ArgAsp                                                                        130                                                                           (2) INFORMATION FOR SEQ ID NO: 159:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 129 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 159:                                    AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        MetValProGlyCysHisGlySerGluProCysIleIleHisArgGly                              202530                                                                        LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        ThrAlaLysIleGluThrLysAlaSerLeuAspGlyLeuGluIleAsp                              505560                                                                        ValProGlyIleAspThrAsnAlaCysHisPheMetLysCysProLeu                              65707580                                                                      ValLysGlyGlnGlnTyrAspAlaLysTyrThrTrpAsnValProLys                              859095                                                                        IleAlaProLysSerGluAsnValValValThrValLysLeuValGly                              100105110                                                                     AspAsnGlyValLeuAlaCysAlaIleAlaThrHisAlaLysIleArg                              115120125                                                                     Asp                                                                           (2) INFORMATION FOR SEQ ID NO: 160:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 92 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 160:                                    AspGlnValAspValLysAspCysAlaAsnSerGluIleLysLysVal                              151015                                                                        MetValProGlyCysHisGlySerGluProCysIleIleHisArgGly                              202530                                                                        LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        ThrAlaLysIleGluIleLysAlaSerLeuAspGlyLeuGluIleAsp                              505560                                                                        ValProGlyIleAspThrAsnAlaCysHisPheMetLysCysProLeu                              65707580                                                                      ValLysGlyGlnGlnTyrAspIleLysTyrThrTrp                                          8590                                                                          (2) INFORMATION FOR SEQ ID NO: 161:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 84 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 161:                                    AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        MetValProGlyCysHisGlySerGluProCysIleIleHisArgGly                              202530                                                                        LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        ThrAlaLysIleGluIleLysAlaSerLeuAspGlyLeuGluIleAsp                              505560                                                                        ValProGlyIleAspThrAsnAlaCysHisPheMetLysCysProLeu                              65707580                                                                      ValLysGlyGln                                                                  (2) INFORMATION FOR SEQ ID NO: 162:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 61 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 162:                                    AspGlnValAspValLysAspCysAlaAsnAsnGluIleLysLysVal                              151015                                                                        MetValProGlyCysHisGlySerGluProCysIleIleHisArgGly                              202530                                                                        LysProPheThrLeuGluAlaLeuPheAspAlaAsnGlnAsnThrLys                              354045                                                                        AlaLysThrGluIleLysAlaSerLeuAspGlyLeuGlu                                       505560                                                                        (2) INFORMATION FOR SEQ ID NO:163:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 32 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:163:                                     LysLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerCysArgArgProAsnAlaGluArg                              202530                                                                        (2) INFORMATION FOR SEQ ID NO:164:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 32 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:164:                                     LysLysGluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyr                              151015                                                                        ArgTyrValAlaArgGluGlnSerCysArgArgProAsnAlaGluLys                              202530                                                                        (2) INFORMATION FOR SEQ ID NO:165:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:165:                                     AspGluGluGlyValValGlnGluSerTyrTyrArgTyrValAlaArg                              151015                                                                        GluGlnSerCysArgArgProAsnAlaGlu                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:166:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:166:                                     AspLysGluGlyValValGlnGluSerTyrTyrArgTyrValAlaArg                              151015                                                                        GluGlnSerCysArgArgProAsnAlaLysGlu                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:167:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:167:                                     AspGluLysGluGlyValValGlnGluSerTyrTyrArgTyrValAla                              151015                                                                        ArgGluGlnSerCysArgArgProAsnAlaAspLysGlu                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO:168:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:168:                                     AspGluGlyIleLysAspLeuAspAlaPheArgHisTyrAspGlyArg                              151015                                                                        ThrIleIleGlnArgAspAsnGlyTyrGlu                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:169:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:169:                                     AspGlnValAspValLysAspCysAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyCysHisGlySerGlu                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO:170:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:170:                                     AspGlnValAspValLysAspCysAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyCysHisGlySerGluGly                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:171:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:171:                                     AspGlnValAspValLysAspCysAlaAsnHisGluIleLysLysVal                              151015                                                                        LeuValProGlyCysHisGlySerGluProLys                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:172:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:172:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaAsp                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:173:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:173:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaLys                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:174:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:174:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAsp                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:175:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:175:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysLys                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:176:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 22 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:176:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLys                                                            20                                                                            (2) INFORMATION FOR SEQ ID NO:177:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:177:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluAsp                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:178:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 21 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:178:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluLys                                                               20                                                                            (2) INFORMATION FOR SEQ ID NO:179:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:179:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGlu                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:180:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:180:                                     AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysAlaGlu                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO:181:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:181:                                     ThrValLysValMetGlyAspAspGlyValLeuAlaCysAlaIleAla                              151015                                                                        ThrHisAlaLysIleArgAsp                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:182:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:182:                                     ThrValLysLeuValGlyAspAspGlyValLeuAlaCysAlaIleAla                              151015                                                                        ThrHisAlaLysIleArgAsp                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO:183:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:183:                                     AspLysThrValLysValMetGlyAspAspGlyValLeuAlaCysAla                              151015                                                                        IleAlaThrHisAlaLysIleArgAspLysGlu                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:184:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:184:                                     AspLysThrValLysLeuValGlyAspAsnGlyValLeuAlaCysAla                              151015                                                                        IleAlaThrHisAlaLysIleArgAspLysGlu                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:185:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 16 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:185:                                     AspLysThrValLysLeuValGlyAspAspGlyValLeuAlaCysAla                              151015                                                                        (2) INFORMATION FOR SEQ ID NO:186:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 16 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:186:                                     AspLysThrValLysLeuValGlyAspAsnGlyValLeuAlaCysAla                              151015                                                                        (2) INFORMATION FOR SEQ ID NO:187:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:187:                                     LysLysThrValLysValMetGlyAspAspGlyValLeuAlaCysAla                              151015                                                                        IleAlaThrHisAlaLysIleArgAspLysLys                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO:188:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:188:                                     AspGluGluThrValLysValMetGlyAspAspGlyValLeuAlaCys                              151015                                                                        AlaIleAlaThrHisAlaLysIleArgAspGluGlu                                          2025                                                                          (2) INFORMATION FOR SEQ ID NO:189:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 30 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:189:                                     AspLysGluLysThrValLysValMetGlyAspAspGlyValLeuAla                              151015                                                                        CysAlaIleAlaThrHisAlaLysIleArgAspLysGluLys                                    202530                                                                        (2) INFORMATION FOR SEQ ID NO:190:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:190:                                     LysLysThrValLysValMetGlyAspAspGlyValLeuAlaCysAla                              151015                                                                        IleAlaThrHisAlaLysLysLys                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO:191:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 26 amino acids                                                    (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:191:                                     AspLysGluLysThrValLysValMetGlyAspAspGlyValLeuAla                              151015                                                                        CysAlaIleAlaThrHisAlaLysLysLys                                                2025                                                                          (2) INFORMATION FOR SEQ ID NO:192:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 549 base pairs                                                    (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA                                                      (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 1..543                                                          (xi) SEQUENCE DESCRIPTION: SEQ ID NO:192:                                     ATTATTGGCATCAAAGATTTAGACGCATTCCGTCATTATGATGGCCGA48                            IleIleGlyIleLysAspLeuAspAlaPheArgHisTyrAspGlyArg                              151015                                                                        ACAATCATTCAACGCGATAATGGTTACCAAACTGTTAAAGTTCTGGGT96                            ThrIleIleGlnArgAspAsnGlyTyrGlnThrValLysValLeuGly                              202530                                                                        GATGATGGTGTTTTGGCCTCTGCTATTGCTACTCATGCTAAAATCCGC144                           AspAspGlyValLeuAlaSerAlaIleAlaThrHisAlaLysIleArg                              354045                                                                        GATGTTGCCGCAACTGAATCAGCTTATTTGGCCTACCGTAACACGTCT192                           AspValAlaAlaThrGluSerAlaTyrLeuAlaTyrArgAsnThrSer                              505560                                                                        TTGGATCTTTCTGAACAGGAACTCGTCGATCAATTGGAAGCCGTTTTC240                           LeuAspLeuSerGluGlnGluLeuValAspGlnLeuGluAlaValPhe                              65707580                                                                      GAAGCCAACCAAAACACAAAAACCGCTAAAATTGAAATCAAAGCCTCA288                           GluAlaAsnGlnAsnThrLysThrAlaLysIleGluIleLysAlaSer                              859095                                                                        ATCGATGGTTTAGAAGTTGAATACATCCAACATAATGGTGTCGTCCAA336                           IleAspGlyLeuGluValGluTyrIleGlnHisAsnGlyValValGln                              100105110                                                                     GAAAGCTACTATCGATACGTTGCACGAGAACAATCATGCCGACGACCA384                           GluSerTyrTyrArgTyrValAlaArgGluGlnSerCysArgArgPro                              115120125                                                                     AATGCACAAGATCAAGTCGATGTCAAAGATTCTGCCAATCATGAAATC432                           AsnAlaGlnAspGlnValAspValLysAspSerAlaAsnHisGluIle                              130135140                                                                     AAAAAAGTTTTGGTACCAGGATCGCATGGTTCAGAACCAAGTATCAAT480                           LysLysValLeuValProGlySerHisGlySerGluProSerIleAsn                              145150155160                                                                  GGAAATGCTCCAGCTGAAATCGATTTGCGACAAATGCGAACTGTCACT528                           GlyAsnAlaProAlaGluIleAspLeuArgGlnMetArgThrValThr                              165170175                                                                     CCCATTCGTATGCAATAATGA549                                                      ProIleArgMetGln                                                               180                                                                           (2) INFORMATION FOR SEQ ID NO:193:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 181 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:193:                                     IleIleGlyIleLysAspLeuAspAlaPheArgHisTyrAspGlyArg                              151015                                                                        ThrIleIleGlnArgAspAsnGlyTyrGlnThrValLysValLeuGly                              202530                                                                        AspAspGlyValLeuAlaSerAlaIleAlaThrHisAlaLysIleArg                              354045                                                                        AspValAlaAlaThrGluSerAlaTyrLeuAlaTyrArgAsnThrSer                              505560                                                                        LeuAspLeuSerGluGlnGluLeuValAspGlnLeuGluAlaValPhe                              65707580                                                                      GluAlaAsnGlnAsnThrLysThrAlaLysIleGluIleLysAlaSer                              859095                                                                        IleAspGlyLeuGluValGluTyrIleGlnHisAsnGlyValValGln                              100105110                                                                     GluSerTyrTyrArgTyrValAlaArgGluGlnSerCysArgArgPro                              115120125                                                                     AsnAlaGlnAspGlnValAspValLysAspSerAlaAsnHisGluIle                              130135140                                                                     LysLysValLeuValProGlySerHisGlySerGluProSerIleAsn                              145150155160                                                                  GlyAsnAlaProAlaGluIleAspLeuArgGlnMetArgThrValThr                              165170175                                                                     ProIleArgMetGln                                                               180                                                                           (2) INFORMATION FOR SEQ ID NO:194:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 549 base pairs                                                    (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA                                                      (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 1..543                                                          (xi) SEQUENCE DESCRIPTION: SEQ ID NO:194:                                     ACTGTTAAAGTTCTGGGTGATGATGGTGTTTTGGCCTCTGCTATTGCT48                            ThrValLysValLeuGlyAspAspGlyValLeuAlaSerAlaIleAla                              151015                                                                        ACTCATGCTAAAATCCGCGATGTTGCCGCAACTGAATCAGCTTATTTG96                            ThrHisAlaLysIleArgAspValAlaAlaThrGluSerAlaTyrLeu                              202530                                                                        GCCTACCGTAACACGTCTTTGGATCTTTCTGAACAGGAACTCGTCGAT144                           AlaTyrArgAsnThrSerLeuAspLeuSerGluGlnGluLeuValAsp                              354045                                                                        GAATACATCCAACATAATGGTGTCGTCCAAGAAAGCTACTATCGATAC192                           GluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyr                              505560                                                                        GTTGCACGAGAACAATCATGCCGACGACCAAATGCACAACAATTGGAA240                           ValAlaArgGluGlnSerCysArgArgProAsnAlaGlnGlnLeuGlu                              65707580                                                                      GCCGTTTTCGAAGCCAACCAAAACACAAAAACGGCTAAAATTGAAATC288                           AlaValPheGluAlaAsnGlnAsnThrLysThrAlaLysIleGluIle                              859095                                                                        AAAGCCTCAATCGATGGTTTAGAAGTTATTATTGGCATCAAAGATTTA336                           LysAlaSerIleAspGlyLeuGluValIleIleGlyIleLysAspLeu                              100105110                                                                     GACGCATTCCGTCATTATGATGGCCGAACAATCATTCAACGCGATAAT384                           AspAlaPheArgHisTyrAspGlyArgThrIleIleGlnArgAspAsn                              115120125                                                                     GGTTACCAAAGTATCAATGGAAATGCTCCAGCTGAAATCGATTTGCGA432                           GlyTyrGlnSerIleAsnGlyAsnAlaProAlaGluIleAspLeuArg                              130135140                                                                     CAAATGCGAACTGTCACTCCCATTCGTATGCAAGATCAAGTCGATGTC480                           GlnMetArgThrValThrProIleArgMetGlnAspGlnValAspVal                              145150155160                                                                  AAAGATTCTGCCAATCATGAAATCAAAAAAGTTTTGGTACCAGGATCG528                           LysAspSerAlaAsnHisGluIleLysLysValLeuValProGlySer                              165170175                                                                     CATGGTTCAGAACCATAATGA549                                                      HisGlySerGluPro                                                               180                                                                           (2) INFORMATION FOR SEQ ID NO:195:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 181 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:195:                                     ThrValLysValLeuGlyAspAspGlyValLeuAlaSerAlaIleAla                              151015                                                                        ThrHisAlaLysIleArgAspValAlaAlaThrGluSerAlaTyrLeu                              202530                                                                        AlaTyrArgAsnThrSerLeuAspLeuSerGluGlnGluLeuValAsp                              354045                                                                        GluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyr                              505560                                                                        ValAlaArgGluGlnSerCysArgArgProAsnAlaGlnGlnLeuGlu                              65707580                                                                      AlaValPheGluAlaAsnGlnAsnThrLysThrAlaLysIleGluIle                              859095                                                                        LysAlaSerIleAspGlyLeuGluValIleIleGlyIleLysAspLeu                              100105110                                                                     AspAlaPheArgHisTyrAspGlyArgThrIleIleGlnArgAspAsn                              115120125                                                                     GlyTyrGlnSerIleAsnGlyAsnAlaProAlaGluIleAspLeuArg                              130135140                                                                     GlnMetArgThrValThrProIleArgMetGlnAspGlnValAspVal                              145150155160                                                                  LysAspSerAlaAsnHisGluIleLysLysValLeuValProGlySer                              165170175                                                                     HisGlySerGluPro                                                               180                                                                           (2) INFORMATION FOR SEQ ID NO:196:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 549 base pairs                                                    (B) TYPE: nucleic acid                                                        (C) STRANDEDNESS: single                                                      (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: cDNA                                                      (ix) FEATURE:                                                                 (A) NAME/KEY: CDS                                                             (B) LOCATION: 1..543                                                          (xi) SEQUENCE DESCRIPTION: SEQ ID NO:196:                                     ACTGTTAAAGTTTTGGGTGATGATGGTGTTTTGGCCTCAGCTATTGCT48                            ThrValLysValLeuGlyAspAspGlyValLeuAlaSerAlaIleAla                              151015                                                                        ACTCATGCTAAAATCCGCGATAGTATCAATGGAAATGCTCCAGCTGAA96                            ThrHisAlaLysIleArgAspSerIleAsnGlyAsnAlaProAlaGlu                              202530                                                                        ATCGATTTGCGACAAATGCGAACTGTCACTCCCATTCGTATGCAAGAA144                           IleAspLeuArgGlnMetArgThrValThrProIleArgMetGlnGlu                              354045                                                                        TACATCCAACATAATGGTGTCGTCCAAGAAAGCTACTATCGATACGTT192                           TyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyrVal                              505560                                                                        GCACGAGAACAATCATGCCGACGACCAAATGCACAAATTATTGGCATC240                           AlaArgGluGlnSerCysArgArgProAsnAlaGlnIleIleGlyIle                              65707580                                                                      AAAGATTTAGACGCATTCCGTCATTATGATGGCCGAACAATCATTCAA288                           LysAspLeuAspAlaPheArgHisTyrAspGlyArgThrIleIleGln                              859095                                                                        CGCGATAATGGTTACCAACAATTGGAAGCCGTTTTCGAAGCCAACCAA336                           ArgAspAsnGlyTyrGlnGlnLeuGluAlaValPheGluAlaAsnGln                              100105110                                                                     AACACAAAAACGGCTAAAATTGAAATCAAAGCCTCAATCGATGGTTTA384                           AsnThrLysThrAlaLysIleGluIleLysAlaSerIleAspGlyLeu                              115120125                                                                     GAAGTTGATCAAGTCGATGTCAAAGATTCAGCCAATCATGAAATCAAA432                           GluValAspGlnValAspValLysAspSerAlaAsnHisGluIleLys                              130135140                                                                     AAAGTTTTGGTACCAGGATCACATGGTTCAGAACCAGTTGCCGCAACT480                           LysValLeuValProGlySerHisGlySerGluProValAlaAlaThr                              145150155160                                                                  GAATCAGCTTATTTGGCCTACCGTAACACGTCTTTGGATCTTTCTGAA528                           GluSerAlaTyrLeuAlaTyrArgAsnThrSerLeuAspLeuSerGlu                              165170175                                                                     CAGGAACTCGTCGATTAGTAG549                                                      GlnGluLeuValAsp                                                               180                                                                           (2) INFORMATION FOR SEQ ID NO:197:                                            (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 181 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:197:                                     ThrValLysValLeuGlyAspAspGlyValLeuAlaSerAlaIleAla                              151015                                                                        ThrHisAlaLysIleArgAspSerIleAsnGlyAsnAlaProAlaGlu                              202530                                                                        IleAspLeuArgGlnMetArgThrValThrProIleArgMetGlnGlu                              354045                                                                        TyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyrVal                              505560                                                                        AlaArgGluGlnSerCysArgArgProAsnAlaGlnIleIleGlyIle                              65707580                                                                      LysAspLeuAspAlaPheArgHisTyrAspGlyArgThrIleIleGln                              859095                                                                        ArgAspAsnGlyTyrGlnGlnLeuGluAlaValPheGluAlaAsnGln                              100105110                                                                     AsnThrLysThrAlaLysIleGluIleLysAlaSerIleAspGlyLeu                              115120125                                                                     GluValAspGlnValAspValLysAspSerAlaAsnHisGluIleLys                              130135140                                                                     LysValLeuValProGlySerHisGlySerGluProValAlaAlaThr                              145150155160                                                                  GluSerAlaTyrLeuAlaTyrArgAsnThrSerLeuAspLeuSerGlu                              165170175                                                                     GlnGluLeuValAsp                                                               180                                                                           (2) INFORMATION FOR SEQ ID NO: 198:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 27 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 198:                                    AspLysGlnLeuGluAlaValPheGluAlaAsnGlnAlaThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaSerIleAspGlu                                             2025                                                                          (2) INFORMATION FOR SEQ ID NO: 199:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 25 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 199:                                    AspGluLysLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaSerAsp                                                   2025                                                                          (2) INFORMATION FOR SEQ ID NO: 200:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 200:                                    AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaAsp                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO: 201:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 24 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 201:                                    AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAlaLys                                                      20                                                                            (2) INFORMATION FOR SEQ ID NO: 202:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 23 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 202:                                    AspLysGluLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThr                              151015                                                                        AlaLysIleGluIleLysAsp                                                         20                                                                            (2) INFORMATION FOR SEQ ID NO: 203:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 16 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 203:                                    AspLysThrAlaLysIleGluIleLysAlaSerIleAspGlyLeuGlu                              151015                                                                        (2) INFORMATION FOR SEQ ID NO: 204:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 15 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 204:                                    LysThrAlaLysIleGluIleLysAlaSerIleAspGlyLeuGlu                                 151015                                                                        (2) INFORMATION FOR SEQ ID NO: 205:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 20 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 205:                                    GluTyrIleGlnHisAsnGlyValValGlnGluSerTyrTyrArgTyr                              151015                                                                        ValAlaArgGlu                                                                  20                                                                            (2) INFORMATION FOR SEQ ID NO: 206:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 29 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 206:                                    HisGlySerGluProCysIleIleHisArgGlyLysProPheGlnLeu                              151015                                                                        GluAlaValPheGluAlaAsnGlnAsnThrLysThrAla                                       2025                                                                          (2) INFORMATION FOR SEQ ID NO: 207:                                           (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 28 amino acids                                                    (B) TYPE: amino acid                                                          (C) STRANDEDNESS:                                                             (D) TOPOLOGY: linear                                                          (ii) MOLECULE TYPE: peptide                                                   (v) FRAGMENT TYPE: internal                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 207:                                    GlnLeuGluAlaValPheGluAlaAsnGlnAsnThrLysThrAlaLys                              151015                                                                        IleGluIleLysAlaSerIleAspGlyLeuGluVal                                          2025                                                                          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We claim:
 1. A therapeutic composition comprising at least one isolatedpeptide selected from the group consisting of: DPI-21.2 (SEQ ID NO:28)and DFI-22.2 (SEQ ID NO:93), and at least one isolated peptide selectedfrom the group consisting of: DPI-23.31 (SEQ ID NO:165), DPI-26.6 (SEQID NO:168), DPII-20.9 (SEQ ID NO:169), DPII-22.14 (SEQ ID NO:180) andDPII-25.15 (SEQ ID NO:188).
 2. A therapeutic composition comprising thefollowing isolated peptides: DPI-21.2 (SEQ ID NO:28), DFI-22.2 (SEQ IDNO:93), DPI-23.31 (SEQ ID NO:165), DPI-26.6 (SEQ ID NO:168), DPII-22.14(SEQ ID NO:180)) and DPII-25.15 (SEQ ID NO: 188).
 3. A method oftreating sensitivity to house dust mite allergen in a mammal comprisingadministering simultaneously or sequentially at least two differentcompositions of claim
 1. 4. A method of treating sensitivity to housedust mite allergen in a mammal comprising administering a therapeuticcomposition of claim
 2. 5. A multipeptide formulation for pharmaceuticaladministration comprising at least one isolated peptide selected fromthe group consisting of: DPI-21.2 (SEQ ID NO:28) and DFI-22.2 (SEQ IDNO:93), and at least one isolated peptide selected from the groupconsisting of: DPI-23.31 (SEQ ID NO:165)), DPI-26.6 (SEQ ID NO: 168),DPII-20.9 (SEQ ID NO:169), DPII-22.14 (SEQ ID NO:180) and DPII-25.15(SEQ ID NO:188), each peptide being soluble and stable at aphysiologically acceptable predetermined pH, and wherein saidformulation comprises a sufficient percentage of T cell epitopes of ahouse dust mite allergen selected from the group consisting of Der p I,Der p II Der f I and Der f II, such that upon administration of themultipeptide formulation to an individual sensitive to house dust mites,T cells of the individual are tolerized to said house dust miteallergen; and a pharmaceutically acceptable carrier.
 6. The multipeptideformulation of claim 5 further comprising EDTA.
 7. The multipeptideformulation of claim 5 further comprising a pharmaceutically acceptablecounter ion.
 8. The multipeptide formulation of claim 5 wherein saidpredetermined pH is a pH in the range of pH 6.0 to pH 8.0.
 9. Themultipeptide of claim 5 wherein said sufficient percentage of T cellactivity is at least 37%.
 10. The multipeptide formulation of claim 5comprising the following isolated peptides: DPI-21.2 (SEQ ID NO:28),DFI-22.2 (SEQ ID NO:93), DPI-23.31 (SEQ ID NO:165), DPI-26.6 (SEQ IDNO:168), DPII-22.14 (SEQ ID NO:180) and DPII-25.15 (SEQ ID NO:188). 11.The multipeptide formulation of claim 5 further comprising the isolatedpeptide, DPII-20.9 (SEQ ID NO:169).
 12. An optimized multipeptideformulation suitable for therapeutic treatment of humans suffering fromallergy to house dust mite comprising:isolated peptides DPI-21.2 (SEQ IDNO:28), DFI-22.2 (SEQ ID NO:93), DPI-23.31 (SEQ ID NO:165), DPI-26.6(SEQ ID NO:168), DPII-22.14 (SEQ ID NO:180) and DPII-25.15 (SEQ IDNO:188), each peptide in a concentration of 0.75 mg per peptide;0.05Msodium phosphate U.S.P.; 5 w/v Mannitol, U.S.P.; 0.1 mg/ml EDTA disodiumdihydrate U.S.P.; and Sterile Water for injection U.S.P;wherein saidformulation has a final pH of 7.2-7.4.
 13. An optimized multipeptideformulation of claim 12 further comprising the isolated peptide,DPII-20.9 (SEQ ID NO:169).
 14. An optimized multipeptide formulationsuitable for therapeutic treatment of humans suffering from allergy tohouse dust mite comprising:isolated peptides DFI-22.2 (SEQ ID NO:93),DPI-23.31 (SEQ ID NO:165), and DPII-22.14 (SEQ ID NO:180), each peptidein a concentration of 0.75 mg per peptide;0.05M sodium phosphate U.S.P.;5% w/v Mannitol, U.S.P.; 0.1 mg/ml EDTA disodium dihydrate U.S.P.; andSterile Water for injection U.S.P;wherein said formulation has a finalpH of 7.0.
 15. An optimized multipeptide formulation suitable fortherapeutic treatment of humans suffering from allergy to house dustmite comprising:isolated peptides DPI-21.2 (SEQ ID NO:28), DPI-26.6 (SEQID NO: 168), and DPII-25.15 (SEQ ID NO:188), each peptide in aconcentration of 0.75 mg per peptide;0.05M sodium phosphate U.S.P.; 5%w/v Mannitol, U.S.P.; 0.1 mg/ml EDTA disodium dihydrate U.S.P.; andSterile Water for injection U.S.P;wherein said formulation has a finalpH of 6.2.
 16. A method of treating sensitivity to house dust miteallergy in an individual comprising administering an optimizedmultipeptide formulation of claim
 12. 17. A method of treatingsensitivity to house dust mite allergy in an individual comprisingadministering simultaneously or sequentially, as a single treatmentepisode, first and second optimized multipeptide formulations whereinsaid first formulation comprises:isolated peptides DFI-22.2 (SEQ IDNO:93, DPI-23.31 (SEQ ID NO:165), and DPII-22.14 (SEQ ID NO:180), eachpeptide in a concentration of 0.75 mg per peptide;0.05M sodium phosphateU.S.P.; 5% w/v Mannitol, U.S.P.; 0.1 mg/ml EDTA disodium dihydrateU.S.P.; and Sterile Water for injection U.S.P; having a final pH of7.0;and wherein said second optimized multipeptide formulationcomprises: isolated peptides DPI-21.2 (SEQ ID NO:28), DPI-26.6 (SEQ IDNO:168, and DPII-25.15 (SEQ ID NO:188), each peptide in a concentrationof 0.75 mg per peptide; 0.05M sodium phosphate U.S.P.; 5% w/v Mannitol,U.S.P.; 0.1 mg/ml EDTA disodium dihydrate U.S.P.; andSterile Water forinjection U.S.P; having a final pH of 6.2.
 18. An isolated peptidehaving an amino acid sequence selected from the group consisting ofDPI-21.2 (SEQ ID NO:28), DFI-22.2 (SEQ ID NO:93), DPI-23.31 (SEQ IDNO:165), DPI-26.6 (SEQ ID NO:168), DPII-20.9 (SEQ ID NO:169), DPII-22.14(SEQ ID NO:180) and DPII-25.15 (SEQ ID NO:188).
 19. A therapeuticcomposition comprising at least one isolated peptide selected from thegroup consisting of: DPI-21.1 (SEQ ID NO:27), DPI-21.2 (SEQ ID NO:28),DFI-22.2 (SEQ ID NO:93), DPI-23.31 (SEQ ID NO:29), DPI-26.6 (SEQ IDNO:168), DPII-22.4 (SEQ ID NO:37), DPII-20.9 (SEQ ID NO:31) andDPII-25.15 (SEQ ID NO:188).
 20. A therapeutic composition of claim 19comprising at least two of said isolated peptides.
 21. A therapeuticcomposition comprising the following isolated peptides: DPI-21.2 (SEQ IDNO:28), DPI-26.6 (SEQ ID NO:168), and DPII-25.15 (SEQ ID NO:50188).